Indole Alkaloid Derivatives Having Opioid Receptor Agonistic Effect, and Therapeutic Compositions and Methods Relating to Same

ABSTRACT

Indole alkaloid derivatives having an opioid receptor agonistic effect, their synthesis, and therapeutic compositions containing these derivatives, and methods of treating conditions with these compounds and therapeutic compositions, are provided.

INTRODUCTION

The section headings used herein are solely for organization purposesand are not to be construed as limiting the subject matter described inany way.

The present invention pertains to indole alkaloid derivatives having anopioid receptor agonistic effect, their synthesis, and therapeuticcompositions containing these derivatives, and additionally methods oftreating conditions with these compounds and therapeutic compositionsare also provided.

BACKGROUND

For the clinical treatment of acute and chronic severe pain, morphine isutilized as a standard analgesic. Morphine-related derivatives have beensynthesized by simplification and introduction of substituents into themorphine structure in order to develop powerful analgesics without sideeffects (Corbett et al., 2006). Analgesics such as fentanyl andbuprenorphine have been consequently derived from morphine. Most of themhave μ-receptor agonist profiles and are used clinically. Despite theirprofound utility in the management of pain, they have undesirable sideeffects such as constipation, respiratory, depression, and developmentof dependence. It is known that h-opioids such as morphine induce notonly potent antinociception but also undesired rewarding effectsfollowing chronic administration in mice. The activation of dopaminergicsystems after systemic administration of a μ-opioid agonist inducesdevelopment of hyperlocomotion and place preference in mice (Matthes etal., 1999). Inhibitory effects on gastrointestinal transit (IGIT), suchas constipation, tend to be a significant problem during administrationof a chronic opioid such as morphine. The dose required for morphine'sanalgesic effect is much higher than that required for its constipatingeffect; thus, when morphine is used for analgesia, constipation is not anegligible issue (Megens et al., 1998).

The traditional Thai herbal medicine Mitragyna speciosa has long beenused in Thailand for its opium- (Burkill, 1935) and coca-like effectsand as a replacement for opium (Grewal, 1932; Suwanlert, 1975). Theleaves of Mitragyna speciosa have been used, and are effective whentaken orally. This medicinal herb contains many indole alkaloids(Takayama 2004). Mitragynine, illustrated below, a main constituent ofthis plant, is an indole alkaloid and structurally different frommorphine, also illustrated below

Studies have been done on the pharmacological activities of mitragynine(Watanabe et al., 1997; Matsumoto et al., 2005) and related alkaloids(Yamamoto et al., 1999; Takayama et al., 2002; Takayama, 2004; Matsumotoet al., 2006a), which have found that these compounds have agonisticeffects on opioid receptors. Recently, studies have been done on theopioid agonistic effects of the constituents of Mitragyna speciosa usingin vitro assays. Among them, 7-hydroxymitragynine, illustrated below,which has a hydroxyl group at the C7 position of mitragynine, producedthe most potent effect, suggesting that the opioid effect of Mitragynaspeciosa is mostly based on the activity of 7-hydroxymitragynine (Horieet al., 2005).

7-Hydroxymitragynine induced a potent antinociceptive effect in mice,and its effect was more potent than those of morphine whensubcutaneously or orally administered and mediated by the μ-opioidreceptor mechanism (Matsumoto et al., 2004; Matsumoto et al., 2006).Furthermore, 7-hydroxymitragynine inhibited gastrointestinal transitless potently than morphine at each equi-antinociceptive dose (Matsumotoet al., 2006). The structural similarities between morphine and7-hydroxymitragynine have been investigated using molecular modelingtechniques (Matsumoto et al., 2005), but could not superimpose all threefunctional groups, i.e., a nitrogen atom, a benzene residue, and anoxygen atom on the benzene ring in the structures of morphine and7-hydroxymitragynine. These functional groups have been considered toplay an important role in producing analgesic activity (Dhawan et al.,1996).

A need has been recognized and solved by the present inventors fordeveloping unique and potent analgesic compounds that can provide painrelief and/or prevention with reduced side effects.

SUMMARY

Uniquely structured indole alkaloid compounds are presented that areuseful for pain treatment and other therapeutic effects with reducedadverse side effects compared to prior alkaloid analgesics such asmorphine and morphine derivatives.

According to various embodiments, indole alkaloid derivatives areprovided having an opioid receptor agonistic effect. The indole alkaloidcompounds of the present teachings have a useful pharmacological profilefor producing potent antinociceptive effects with fewer rewardingeffects compared with μ-agonists in general, and weaker adverse sideeffects, such as in terms of IGIT, than morphine in particular.

According to various embodiments, indole alkaloid compound having anopioid receptor agonistic effect is provided having the formula (a):

wherein X represents hydrogen nor a halogen atom, Q and A eachindependently represents hydrogen, alkyl, or aryl, E represents ahydrogen or a hydroxyl group, m is 0 or 1, n is 0 or 1, q is 0 or 1, R₁and R₂ independently are hydrogen, hydroxy, alkoxy, aryloxy, alkyl,aryl, aralkyl, alkaryl, alkyl amide, amino, alkylamino, halogen,fluorinated alkyl, fluorinated alkoxy, nitro group, or cyano group, R₃represents an ester group or carboxyl group, wherein the dashed lineseach indicates an optional double bond with the provisos that when q is0 and X is halogen then a 2,7 double bond is present and when m is 0, qis 1 and X is halogen then a 1,2 double bond is present. In anotherembodiment, when m, q and n each are 1, then no 1,2 or 2,7 positiondouble bond is present in the indole moiety of the compound. In afurther embodiment of formula (a), R₁ and R₂ independently are C₁-C₆alkoxy, and R₃ represents a C₁-C₆ alkyl carboxylic acid ester group, andQ and A (if present) are hydrogen, and the other groups and mentionedbonds are the same as previously defined.

According to other various embodiments, a C10-halogenated indolealkaloid compound having an opioid receptor agonistic effect is providedthat is selected from the formulae:

where R is a halogen atom, and R₁, R₂ and R₃ have the same respectivemeanings as defined above for formula (a).

According to other various embodiments, the C10-halogenated indolealkaloid compound having an opioid receptor agonistic effect is selectedfrom the formulae:

wherein the R is fluorine, chlorine, bromine, or iodine.

According to other various embodiments, the C10-halogenated indolealkaloid compound is C10-halogenated mitragynine,7-hydroxy-C10-halogenatedmitragynine, C₁₋₁₀-halogenated2,3-dimethylindole, C10-halogenated tetrahydrocarbazole, C10-halogenatedindoloquinolizidine corynantheol, C10-halogenated dihydrocorynantheol,or C10-halogenated yohimbine, singly or in any combination thereof.According to further embodiments, the compound is 10-halomitragynine,7-hydroxy-10-halomitragynine, singly or in a combination thereof.

According to other various embodiments, an indole alkaloid compoundhaving an opioid receptor agonistic effect is provided having theformula:

where X, R₁, R₂ and R₃ have the same respective meanings as definedabove for formula (a).

According to other various embodiments, the indole alkaloid compoundhaving an opioid receptor agonistic effect has the formula:

where X has the same meaning as defined above for formula (a).

According to other various other embodiments, a pharmaceuticalcomposition is provided comprising a therapeutic amount of at least oneindole alkaloid derivative compound having an opioid receptor agonisticeffect or a pharmaceutically acceptable salt thereof of the presentinvention. The indole alkaloid derivative compound contained in thesepharmaceutical compositions can be, for example, selected from amongstone of any of formulae (a), 1, 2, I, II, 1a, and Ia as set forth herein.These indole alkaloid derivative compounds may be used alone or incombinations thereof in a pharmaceutical composition. The pharmaceuticalcomposition can further comprise a pharmaceutically acceptable carrier.The pharmaceutical composition can be provided in a pharmaceuticallydeliverable form, such as solutions, suspensions, emulsions, tablets,pills, pellets, granules, lozenges, capsules, capsules containingliquids, powders, sustained-release formulations, syrups, elixirs,creams, gels, suppositories, emulsions, aerosols, or sprays, In variousembodiments, the indole alkaloid derivative (as an active ingredient)can be compounded in salt form into tablets or pills and the like, oralternatively can be dissolved in solution. The pharmaceuticalcomposition can contain, for example, 0.1 wt % to 100 wt %, particularlyabout 1 wt % to about 90 wt %, more particularly from about 5 wt % toabout 80 wt %, of the indole alkaloid derivative compound or apharmaceutically acceptable salt thereof. Other wt % can be from 10 wt %to 75 wt %; from 15 wt % to 60 wt %, from 20 wt % to 50 wt % of theactive ingredient.

According to other various embodiments, a method is provided foreliciting a therapeutic effect in a patient (e.g., human, animal) inneed thereof, comprising the step of administering to the patient aneffective dose of an indole alkaloid derivative compound having anopioid receptor agonistic effect or a pharmaceutically acceptable saltthereof of the present invention. The indole alkaloid derivativecompound can be selected, for example, from amongst any one of formula(a), 1, 2, I, II, 1a or Ia as set forth herein, wherein X or R, asapplicable, is fluorine, chlorine, bromine, or iodine.

According to various embodiments, the therapeutic effect achieved by themethod comprises an analgesic effect, although not limited thereto. Invarious embodiments, the indole alkaloid compounds of the presentteachings can be used to treat or prevent acute or chronic pain. Thestep of administering can be performed, for example, by a delivery routeselected from oral, transdermal, intramuscular, intravenous, inhalation,injection, infusion, or suppository. The patient can be, for example, ananimal, such as a mammal, although not limited thereto. According toparticular embodiments, the patient is a human. Where the patient is ahuman and the therapeutic effect is pain treatment, the effective totaldaily dosage of the indole alkaloid derivative compound can range, forexample, from about 0.1 mg to about 1,000 mg active compound/kg bodyweight of the patient. Other dosages are possible.

According to various embodiments, a method is provided for synthesizinga 10-halo-substituted indole alkaloid derivative compound, such as thoseexemplified in formulae 1, 2, I and II, wherein a Corynanthe-type indolealkaloid is reacted with hypervalent iodine in the presence of ethyleneglycol effective to provide a 2,3-ethylene glycol bridged indolineadduct, effective to mask a 2,3-π bond of an indole nucleus of theadduct. A halogen atom is introduced at a C10 position of the adduct viaelectrophilic aromatic substitution, providing a C10-halogenated adductderivative. The C10-halogenated adduct derivative is converted into acorresponding C10-halogenated mitragynine derivative by reductionreaction effective to eliminate the ethylene glycol bridge. Optionally,the C10-halogenated mitragynine derivative is further converted into acorresponding 7-hydroxy-10-halomitragynine derivative by oxidationreaction.

According to other various embodiments, a method is provided forsynthesizing an indole alkaloid derivative compound, such as thoseexemplified in formulae 1a and Ia, wherein 7-hydroxymitragynine or7-hydroxy-10-halomitragynine is reacted with a reducing agent, such asNaBH₄, in a solvent such as MeOH, to yield compounds of formulae 1a andIa.

The reaction product obtained by the methods according to variousembodiments of the present invention can contain the indole alkaloidderivative compound in a concentration, for example, of from about 5 wt% to 100 wt %, particularly from about 10 wt % to about 99 wt %, about20 wt % to about 99 wt %, and 50 wt % to about 99 wt %.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows dose-response curves for analgesic effect oforally-administered compound 1, compound 11, compound 26, compound 27and morphine in tail-flick test in mice, as discussed in Example 2herein. Analgesia was quantified using the percentage of maximumpossible effect (% MPE) and calculated as: % MPE=[(test latency−pre-druglatency)/(cut-off time−pre-drug latency)]×100. A cut-off time of 10 swas used to prevent tissue damage. Each value represents mean±S.E.M. ofdata obtained from 7 to 9 mice.

FIG. 2 a)-c) shows effects of opioid receptor antagonists on analgesiainduced by subcutaneous (s.c.) administration of compound 11, compound26 and compound 27, as discussed in Example 2 herein. The analgesiceffects were determined in the mouse tail-flick test after s.c.administration of the following antagonists: β-funaltrexamine (β-FNA, 40mg/kg), naltrindole (NTI, 3 mg/kg), and nor-binaltorphimine (norBNI, 20mg/kg). Measurements were performed 15 min after s.c. administration ofcompound 11, compound 26 and compound 27. The doses of compound 11 (1mg/kg, s.c.), compound 26 (0.5 mg/kg, s.c.) and compound 27 (0.1 mg/kg,s.c.) that produce a response of 80-90% were chosen to detect the effectof each antagonist easily. Each value represents mean±S.E.M. of 8 or 9mice. The # denotes values that were significantly different fromvehicle-treated mice by Student's t-test (##, P<0.01). The asterisk (*)denotes values that were significantly different from mice treated withcompound 11, compound 26 and compound 27 alone in one-way analysis ofvariance followed by Bonferroni multiple comparison test (*, P<0.05, **,P<0.01).

FIG. 3 shows dose-response curves of inhibitory effect ongastrointestinal transit of orally-administered compound 1, compound 11,and morphine in mice, as discussed in Example 2 herein. The inhibitionof gastrointestinal transit (%) was calculated as: Inhibition ofgastrointestinal transit (%)=[(saline or vehicle GIT−drug GIT)/(salineor vehicle GIT)]×100. Each value represents mean±S.E.M. of 7 or 8 mice.

FIG. 4 a is the 1H-NMR for Ethylene Glycol-Bridged Compound 3 and FIG. 4b is the 13C-NMR for Ethylene Glycol-Bridge Compound 3.

FIG. 5 a is the 1H-NMR for Compound 4 and FIG. 5 b is the 13C-NMR forCompound 4.

FIG. 6 a is the 1H-NMR for Compound 5 and FIG. 6 b is the 13C-NMR forCompound 5.

FIG. 7 a is the 1H-NMR for Compound 6 and FIG. 7 b is the 13C-NMR forCompound 6.

FIG. 8 a is the 1H-NMR for Compound 11 and FIG. 8 b is the 13C-NMR forCompound 11.

FIG. 9 a is the 1H-NMR for Compound 17 and FIG. 9 b is the 13C-NMR forCompound 17.

FIG. 10 a is the 1H-NMR for Compound 22 and FIG. 10 b is the 13C-NMR forCompound 22.

DESCRIPTION OF VARIOUS EMBODIMENTS

It is to be understood that the following descriptions are exemplary andexplanatory only. The accompanying drawings are incorporated in andconstitute a part of this application and illustrate several exemplaryembodiments with the description. Reference will now be made to variousembodiments, examples of which are illustrated in the accompanyingdrawings.

For purposes of this application, the following definitions apply.

“Halogen” or “-Halo” means fluorine, chlorine, bromine, or iodine.

The phrase “pharmaceutically acceptable salt,” as used herein, can be asalt formed, for example, from an acid and a basic functional group,such as a nitrogen group, of the indole alkaloid compound. Illustrativesalts include, but are not limited, to sulfate, citrate, acetate,oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acidphosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate,oleate, tannate, pantothenate, bitartrate, ascorbate, succinate,maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate,formate, benzoate, glutamate, methanesulfonate, ethanesulfonate,benzenesulfonate, p-toluenesulfonate, pamoate salts, or hydrochloridesalts thereof.

The phrases “therapeutic amount,” “therapeutic dose,” “effectiveamount,” “effective dose,” and the like, when used in connection with anindole alkaloid compound means an amount effective for treating and/orpreventing a condition.

The phrases “treatment of,” “treating,” and the like include theamelioration or cessation of a condition, or a symptom thereof.

The phrases “prevention of,” “preventing,” and the like include theavoidance of the onset of a condition, or a symptom thereof.

The term “animal,” for purposes of treatment, refers to a mammal, bird,reptile or fish. Representative animals include, for example, a monkey,chimpanzee, baboon, cow, buffalo, horse, sheep, pig, chicken, turkey,duck, cat, dog, rabbit, mouse, rat, hamster, guinea pig, and human.

The term “mammal” for purposes of treatment refers to any animalclassified as a mammal, including humans, domestic and farm animals, andzoo, sport, or pet animals.

Indole Alkaloid Derivative Structures

Uniquely structured indole alkaloid compounds are provided in accordancewith the present teachings that can be used for pain treatment and othertherapeutic effects with reduced adverse side effects compared to prioralkaloid analgesics such as morphine and morphine derivatives.

According to various embodiments, indole alkaloid compound having anopioid receptor agonistic effect is provided having the formula (a)indicated below:

wherein:X represents halogen atom or hydrogen,Q and A each independently represents hydrogen, alkyl, or aryl,E represents a hydrogen or hydroxyl group,m is 0 or 1,n is 0 or 1,q is 0 or 1,R₁ and R₂ independently are hydrogen, hydroxy, alkoxy, aryloxy, alkyl,aryl, aralkyl, alkaryl, alkyl amide, amino, alkylamino, halogen,fluorinated alkyl, fluorinated alkoxy, nitro group, or cyano group,R₃ represents an ester group or carboxyl group,the dashed lines each indicates an optional double bond with provisosthat when q is 0 and X is halogen then a 2,7 double bond is present, andwhen m is 0, q is 1 and X is halogen then a 1,2 double bond is present.In another embodiment, when m, q and n each is 1, then no 1,2 or 2,7position double bond is present in the indole moiety of the compound,

In a further embodiment of the compounds of formula (a), R₁ and R₂independently are C₁-C₆ alkoxy, R₃ represents a C₁-C₆ alkyl carboxylicacid ester group, and Q and A (if present) are hydrogen, and the othergroups and mentioned bonds are the same as previously defined. The C₁-C₆alkoxy groups can be, e.g., methoxy, ethoxy, propoxy, butoxy, and thelike), and C₁-C₆ alkyl carboxylic acid ester group can be, e.g.,acetate, ethanoate, butanoate, hexanoate, and the like, R₁ and R₂ can bethe same or different. R₁, R₂, and R₃ can be substituted orunsubstituted. Any C₃ or greater alkyl portions of these groups can belinear or branched.

According to other various embodiments, a C10-halogenated indolealkaloid compound having opioid receptor agonistic effect is providedthat is selected from the formulae:

where R has the same meaning as X as defined above, R₁, R₂ and R₃ havethe same respective meanings as defined above.

According to other various embodiments, the C10-halogenated indolealkaloid compound having opioid receptor agonistic effect is selectedfrom the formulae:

wherein the R is fluorine, chlorine, bromine, or iodine.

According to other various embodiments, the C10-halogenated indolealkaloid compound is C10-halogenated mitragynine,7-hydroxy-C10-halogenatedmitragynine, C10-halogenated2,3-dimethylindole, C10-halogenated tetrahydrocarbazole, C10-halogenatedindoloquinolizidine corynantheol, C10-halogenated dihydrocorynantheol,or C10-halogenated yohimbine, singly or in any combination thereof.

According to further embodiments, the compound is 10-halomitragynine,7-hydroxy-10-halomitragynine, singly or in a combination thereof.Specific therapeutic compounds according to embodiments of the presentteachings include, for example, 7-hydroxy-10-fluoromitragynine,7-hydroxy-10-chloromitragynine, 7-hydroxy-10-bromomitragynine,10-fluoromitragynine, 10-chloromitragynine, or 10-bromomitragynine.

According to other various embodiments, an indole alkaloid compoundhaving an opioid receptor agonistic effect is provided having theformula:

where X, R₁, R₂ and R₃ have the same respective meanings as definedabove for formula (a).

According to other various embodiments, the indole alkaloid compoundhaving an opioid receptor agonistic effect has the formula:

where X has the same meaning as defined above for formula (a).

According to various embodiments, the indole alkaloid compounds of thepresent teachings have a useful pharmacological profile for producingpotent antinociceptive effects with fewer rewarding effects comparedwith μ-agonists in general and weaker adverse side effects than morphinein particular. According to various embodiments, indole alkaloidcompounds of the present teachings can inhibit gastrointestinal transitless potently than morphine at each equi-antinociceptive dose. Compoundsof formula 1a (and Ia) have shown oral analgesic activity that isgreater than about 200 times or more potent than that of morphine. Theyalso exhibit potent in vitro activity relative to other mitragyninederivatives.

Although not desiring to be bound to theory, it is thought that theindole alkaloid compounds of the present teachings are unique dualacting μ- and κ-opioid agonists, which can produce strongerantinociceptive effects and weaker adverse effects than morphine. Asdiscussed previously, μ-opioids induce potent antinociception, but theyalso induce psychological dependence during chronic administration.Activation of dopaminergic systems after administration of the μ-opioidagonist induces the development of rewarding effects. In contrast,κ-opioid receptors negatively modulate the activity of dopaminergicneurons and inhibit the rewarding effects mediated by μ-opioid receptors(Narita et al., 2001). Therefore, it is hypothesized that a dual actingμ- and κ-opioid agonist will induce potent antinociceptive effects andfewer rewarding effects than μ agonists such as morphine. Theinvolvement of μ- and κ-opioid receptor mechanisms in the opioidagonistic effects of indole alkaloid compounds of the present teachingsis possible. The affinities of indole alkaloid compounds of embodimentsof the present teachings for the three opioid receptor types of μ-, δ-,and κ-opioid receptors can be determined by evaluating the inhibition ofbinding of ligands to μ-, δ-, and κ-opioid receptors.

Synthesis Methods

Synthesis of C10-Halogenated Indole Alkaloid Compounds of Formula 1.

Synthesis methods that can be utilized or adapted in preparingC10-halogenated indole alkaloid compounds of embodiments of the presentteachings are described, for example, in Takayama, H., et al. (2006) andTakayama, H., et al. (2002).

In one embodiment of the present invention, the inventors have found anew method to protect the 2,3-κ of indole alkaloids, which has beenapplied to the preparation of derivatives having various substituents atthe C10-position in Corynanthe-type indole alkaloids such ashydroxymitragynine or its parent compound.

The present inventors have determined that attempts at the directintroduction of electrophilic substituents on the benzene ring in7-hydroxymitragynine or its parent compound are not successful. Thepresent investigators have devised a method to protect the 2,3-π ofindole alkaloids, producing the aniline structure that acts as areactive aromatic compound toward various electrophiles. According toone embodiment, the general synthesis scheme is shown in Scheme A.

With reference to Scheme A, in accordance with embodiments of thepresent teachings, treatment of indole alkaloids with hypervalent iodinein the presence of ethylene glycol provides 2,3-ethylene glycol bridgedadducts that can be converted into the original indoles under mildreductive conditions. This procedure, which involves masking of thereactivity of the indole nucleus at the β-position, is used for themodification of the benzene ring of the indoline derivative and issupplied to the preparation of potent opioid receptor agonists with theCorynanthe skeleton. The purity of the reaction product compounds can bechecked, for example, by high-performance liquid chromatography and¹H-nuclear magnetic resonance (500 MHz) analysis.

Synthesis of Indole Alkaloid Compounds of Formula 1a.

A method also is provided for synthesizing an indole alkaloid derivativecompound, such as those exemplified in formulae 1a and Ia, wherein7-hydroxymitragynine or 7-hydroxy-10-halomitragynine, or a similarindole alkaloid, is reacted with an indole reducing agent, such asNaBH₄, in a solvent such as MeOH, to yield compounds of formulae 1a andIa. According to one embodiment thereof, the general synthesis scheme isshown in Scheme B.

where X is a halogen atom or a hydrogen atom depending on the startingcompound.

The reaction product obtained by methods according to embodiments of thepresent teachings, such as illustrated above, can contain the indolealkaloid derivative compound in a concentration, for example, of fromabout 5 wt % to 100 wt %, particularly from about 10 wt % to about 99 wt%, from about 20 wt % to about 99 wt %, or from 50 wt % to about 99 wt%.

Therapeutic/Prophylactic Administration and Compositions

Pharmaceutical compositions according to embodiments of the presentteachings can contain, for example, from 0.1 wt % to 100 wt %,particularly from about 1 wt % to about 90 wt %, more particularly fromabout 5 wt % to about 80 wt %, of the indole alkaloid derivativecompound or a pharmaceutically acceptable salt thereof, which amountscan vary depending on factors such as the overall formulation and formthereof, intended treatment, and patient details at hand, and the like.

Due to their activity, the indole alkaloid compounds of embodimentsaccording to the present teachings are advantageously useful, forexample, in veterinary and human medicine. As described above, theindole alkaloid compounds are useful for treating or preventing acondition in an animal in need thereof. When administered to an animal,the indole alkaloid compounds are administered as a component of acomposition that comprises any pharmaceutically acceptable carrier orexcipient. The present compositions, which comprise at least one indolealkaloid compound, can be administered orally. The indole alkaloidcompounds of the invention can also be administered by any otherconvenient route, for example, by infusion or bolus injection, byabsorption through epithelial or mucocutaneous linings (e.g., oral,rectal, and intestinal mucosa, etc.) and can be administered togetherwith another biologically active agent. Administration can be systemicor local. Various delivery systems are known, e.g., encapsulation inliposomes, microparticles, microcapsules, capsules, etc., and can beused to administer the indole alkaloid compounds.

Methods of administration include, but are not limited to, intradermal,intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal,epidural, oral, sublingual, intracerebral, intravaginal, transdermal,rectal, by inhalation, or topical, particularly to the ears, nose, eyes,or skin. The mode of administration is left to the discretion of thepractitioner. In most instances, administration will result in therelease of the C10-halogenated indole alkaloid compounds into thebloodstream.

In specific embodiments, it can be desirable to administer the indolealkaloid compounds locally. This can be achieved, for example, and notby way of limitation, by local infusion during surgery, topicalapplication, e.g., in conjunction with a wound dressing after surgery,by injection, by means of a catheter, by means of a suppository orenema, or by means of an implant, said implant being of a porous,non-porous, or gelatinous material, including membranes, such assialastic membranes, or fibers.

In certain embodiments, it can be desirable to introduce the indolealkaloid compounds into the central nervous system or gastrointestinaltract by any suitable route, including intraventricular, intrathecal,and epidural injection, and enema. Intraventricular injection can befacilitated by an intraventricular catheter, for example, attached to areservoir, such as an Ommaya reservoir. In certain embodiments, it canbe desirable to administer the indole alkaloid compounds by subcutaneousadministration, such as, for example, as dissolved in phosphate-bufferedsaline (pH 5.3-5.5).

Pulmonary administration can also be employed, e.g., by use of aninhaler or nebulizer, and formulation with an aerosolizing agent, or viaperfusion in a fluorocarbon or synthetic pulmonary surfactant. Incertain embodiments, the indole alkaloid compounds can be formulated asa suppository, with traditional binders and excipients such astriglycerides.

In another embodiment, the indole alkaloid compounds can be delivered ina vesicle, in particular a liposome. In yet another embodiment, theindole alkaloid compounds can be delivered in a controlled-releasesystem or sustained-release system. In one embodiment, a pump can beused. In another embodiment, polymeric materials can be used. In yetanother embodiment, a controlled- or sustained-release system can beplaced in proximity of a target of the indole alkaloid compounds, e.g.,the spinal column, brain, or gastrointestinal tract, thus requiring onlya fraction of the systemic dose.

The present compositions can optionally comprise a suitable amount of apharmaceutically acceptable excipient so as to provide the form forproper administration to the animal. Such a pharmaceutical excipient canbe a liquid, such as water or an oil, including those of petroleum,animal, vegetable, or synthetic origin, such as peanut oil, soybean oil,mineral oil, sesame oil and the like. The pharmaceutical excipient canbe saline, gum acacia, gelatin, starch paste, talc, keratin, colloidalsilica, urea and the like. In addition, auxiliary, stabilizing,thickening, lubricating, and coloring agents can be used. In oneembodiment, the pharmaceutically acceptable excipient is sterile whenadministered to an animal. Water is a particularly useful excipient whenthe indole alkaloid compound is administered intravenously. Salinesolutions and aqueous dextrose and glycerol solutions can also beemployed as liquid excipients, particularly for injectable solutions.Suitable pharmaceutical excipients also include starch, glucose,lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodiumstearate, glycerol monostearate, talc, sodium chloride, dried skim milk,glycerol, propylene, glycol, water, ethanol and the like. The presentcompositions, if desired, can also contain minor amounts of wetting oremulsifying agents, or pH buffering agents.

The present compositions can take the form of solutions, suspensions,emulsions, tablets, pills, pellets, capsules, capsules containingliquids, powders, sustained-release formulations, suppositories,emulsions, aerosols, sprays, suspensions, or any other form suitable foruse. In one embodiment, the composition is in the form of a capsule.Other examples of suitable pharmaceutical excipients are described inRemington's Pharmaceutical Sciences, incorporated herein by reference.In one embodiment, the indole alkaloid compounds are formulated inaccordance with routine procedures as a composition adapted for oraladministration to human beings. Compositions for oral delivery can be inthe form of tablets, lozenges, aqueous or oily suspensions, granules,powders, emulsions, capsules, syrups, or elixirs, for example. Orallyadministered compositions can contain one or more agents, for example,sweetening agents such as fructose, aspartame or saccharin; flavoringagents such as peppermint, oil of wintergreen, or cherry; coloringagents; and preserving agents, to provide a pharmaceutically palatablepreparation. Moreover, where in tablet or pill form, the compositionscan be coated to delay disintegration and absorption in thegastrointestinal tract thereby providing a sustained action over anextended period of time. Selectively permeable membranes surrounding anosmotically active driving compound are also suitable for orallyadministered compositions. In these latter platforms, fluid from theenvironment surrounding the capsule is imbibed by the driving compound,which swells to displace the agent or agent composition through anaperture. These delivery platforms can provide an essentially zero orderdelivery profile as opposed to the spiked or bolus profiles of immediaterelease formulations. A time-delay material such as glycerol(mono)stearate can also be used. Oral compositions can include standardexcipients such as mannitol, lactose, starch, magnesium stearate, sodiumsaccharin, cellulose, or magnesium carbonate. In one embodiment, theexcipients are of pharmaceutical grade.

In another embodiment, the indole alkaloid compounds can be formulatedfor intravenous administration. Typically, compositions for intravenousadministration can comprise a sterile isotonic aqueous buffer. Wherenecessary, the compositions can also include a solubilizing agent.Generally, the ingredients are supplied either separately or mixedtogether in unit dosage form, for example, as a dry lyophilized powderor water free concentrate in a hermetically sealed container such as anampule or sachette indicating the quantity of active agent. Where theindole alkaloid compounds are to be administered by infusion, they canbe dispensed, for example, with an infusion bottle containing sterilepharmaceutical grade water or saline. Where the indole alkaloidcompounds are administered by injection, an ampule of sterile water forinjection or saline can be provided so that the ingredients can be mixedprior to administration.

The amount of the indole alkaloid compounds that is effective for thetreatment or prevention of a condition can be determined by standardclinical techniques. In addition, in vitro or in vivo assays canoptionally be employed to help identify optimal dosage ranges. Whenadministered for the treatment and/or prevention of a particulardisorder and/or disease state, it is understood that the effectivedosage may vary depending upon the particular compound used, the mode ofadministration, the condition, and severity thereof, of the conditionbeing treated, as well as the various physical factors related to thepatient being treated, such that the precise dosing regimen to beemployed can also depend on these factors and the judgment of a healthcare practitioner treating the patient. Suitable effective dosageamounts, for example, can range from about 0.1 mg/kg body weight/day toabout 1,000 mg/kg body weight/day, particularly from about 1 mg/kg bodyweight/day to about 200 mg/kg body weight/day, and more particularlyfrom about 2 mg/kg body weight/day to about 100 mg/kg body weight/day.The dosage amount can be administered in a single dosage treatment perday or as divided into separate smaller dosage treatments per day,depending on the treatment circumstances. For example, in paintreatment, from about 5 to about 30 mg can be administered to a patientin need thereof every four hours or other dosage and/or interval, asneeded. The indole alkaloid compounds also can be administered as anaqueous solution or other solution containing the compound or saltthereof, and such as in a concentration of from about 1 mg to about 25mg per mL or other concentration.

The opioid component of the present methods and compositions may furtherinclude one or more other active ingredients that may be conventionallyemployed in analgesic and/or cough-cold-antitussive combinationproducts. Such conventional ingredients include, for example, aspirin,acetaminophen, phenylpropanolamine, phenylephrine, chlorpheniramine,caffeine, and/or guaifenesin, and the like. Typical or conventionalingredients that may be included in the opioid component are described,for example, in the Physicians' Desk Reference, the disclosure of whichis hereby incorporated herein by reference, in its entirety.

In addition, the opioid component may further include one or morecompounds that may be designed to enhance the analgesic potency of theopioid and/or to reduce analgesic tolerance development. Although thecompounds of the present invention may be administered as the purechemicals, the active ingredient can be administered as a pharmaceuticalcomposition. The invention thus further provides a pharmaceuticalcomposition comprising an effective amount of one or more of thecompounds of the invention, preferably one or more compounds describedherein, together with one or more pharmaceutically acceptable carrierstherefore and, optionally, other therapeutic and/or prophylacticingredients. The carrier(s) must be acceptable in the sense of beingcompatible with the other ingredients of the composition and notdeleterious to the recipient thereof. The compounds of the invention maybe administered in an effective amount by any of the conventionaltechniques well-established in the medical field. The compounds employedin the methods of the present invention including, for example, opioidsand the compounds of the invention, or a pharmaceutically acceptablesalt thereof, may be administered by any means that results in thecontact of the active agent(s) with the relevant site or site(s) ofaction in the body of a patient. The compounds may be administered byany conventional means available for use in conjunction withpharmaceuticals, either as individual therapeutic agents or in acombination of therapeutic agents. For example, they may be administeredas the sole active agents in a pharmaceutical composition, or they canbe used in combination with other therapeutically active ingredients.

The present invention will be further clarified by the followingexamples, which are intended to be purely exemplary of the presentinvention. Unless otherwise indicated, all numbers expressingquantities, percentages or proportions, and other numerical values usedin the specification and claims, are to be understood as being weightpercentages unless indicated to the contrary.

EXAMPLES Example 1 C10-Substituted Indole Alkaloid Compounds Synthesisof C10-Substituted Indole Alkaloid Compounds

Mitragynine (compound 2) was treated with 1 equivalent of phenyliodinebis (trifluoroacetate)(PIFA) in the presence of ethylene glycol (EG) inMeCN at 0° C., and a 2,3-ethylene glycol bridged indoline derivative wasobtained in quantitative yield. The structure of the adduct includingthe stereochemistry was determined from spectroscopic data, as shown inScheme 1.

Indoline compound 3 could be converted into the starting indole,mitragynine, in almost quantitative yield upon reduction with Na CNBH₃in AcOH at room temperature, followed by heating at 90° C. afteraddition of MeOH. Indoline 3 was put to practice use for the preparationof several benzene-substituted derivatives for the study of opioidreceptor ligands, as described below.

Using other indole alkaloids, the present investigators examined thegenerality of the newly developed method to mask the pyrrole moiety inthe indole nucleus. Among the tested compounds 2,3-dimethylindole,tetaracarbazole, indoloquinolizidine corynantheeol, dihydrocorynantheol,and yohimbine, the corresponding EG adducts (4-9), were obtained inmoderate yields. However, the best results (the yields are shown inScheme 1a) were obtained when NH₄Cl was added to the reaction mixture(see Supporting Information).

In the case of reserpine, it was found that phenyliodine diacetate(PIDA) was a more suitable reagent than PIFA for the formation of theEG-bridged adduct (10), which was also useful as a starting material forthe preparation of various kinds of A-ring-modified reserpine analogues.

Using EG adduct 3 derived from mitragynine (2), various kinds ofsubstituents were introduced onto the benzene ring as shown in Scheme 2.

Treatment of compound 3 with N-fluoro-2,6-dichloropyridinium triflate(FP-T800) gave compound 11 fluorinated at the C-10 position in 53%yield. Exposure of compound 3 to NCS in AcOH afforded two chlorinatedderivatives 12a (10-Chloro) and 12b (12-Chloro) in 88% and 11% yieldsrespectively. Using NBS in DMF, 10-bromo and 12-bromo derivatives (13aand 13b) were obtained in 75% and 24% yields, respectively. To introducea nitro group, for comparison sake, a combination of CAN andconcentrated H₂SO₄ in DCM was used to give 14a in 52% yield togetherwith its 12-isomer (14b) in 21% yield. 10-Methoxy derivative 15 wasprepared in 64% yield by treatment of compound 3 with IBDA in MeOH,followed by the reduction of the resulting iminoquinone intermediate(see Supporting Information) with Zn in MeOH.

The C10-substituted derivative thus obtained as a major product of eachelectrophilic aromatic substitution reaction were converted into theirindole derivatives in good yields by reduction with NaCNBH₃ in AcOH asdescribed above (conversion from compound 3 into compound 2). However,in the case of nitro derivative 14a, a two-step procedure was used;i.e., compound 14a was treated with TBSOTf in the presence of2,6-lutadine and the resultant indolenine derivative 16 obtained in 87%yield was reduced with NaCNBH₃ to give the indole derivative 20 in 94%yield (Scheme 3). The thus obtained indole derivatives were,respectively, converted into 7-hydroxyindolenine derivatives (22-25) byoxidation with PIFA in aqueous MeCN.

Additional details on the experimental procedures used for thepreparation of compounds 2-25, and copies of ¹H and ¹³C NMR spectraldata for compounds 3-6, 11, 17 and 22 are provided in the SupportingInformation included herewith.

Pharmacological Evaluation

The series of C10-substituted mitragynine derivatives obtained by theabove reactions were subjected to pharmacological evaluation. The opioidagonistic effect was evaluated in an experiment involving twitchcontraction induced by electrical stimulation of guinea pig ileum. Thisexperiment is generally used to study opioid analgesics. Additionaldetails on the experimental procedures used for this evaluation areprovided in the Supporting Information. The results are shown in TableA.

TABLE A Opioid Effects of Mitragynine Derivatives on Twitch ContractionInduced by Electrical Stimulation in Guinea Pig Ileum^(a) pD2 valueRelative Maximum Inhibitory Compound (−log M) Potency (%) inhibition (%)activity (%) Morphine 7.15 ± 0.05 100 87.2 ± 1.8 100 Ethylene GlycolBridged Derivatives  3 7.70 ± 0.10 354  35.0 ± 11.0 40 11 8.40 ± 0.021778 83.4 ± 3.2 96 12a 7.61 ± 0.17 288 48.1 ± 9.3 55 14a 7.88 ± 0.18 53765.0 ± 4.3 75 Mitragynine Derivatives  2 6.50 ± 0.06 22 72.0 ± 5.0 837-Hydroxyindolenine Derivatives  1 7.78 ± 0.10 426 90.8 ± 3.4 104 227.87 ± 0.04 524 82.5 ± 1.8 95 23 7.53 ± 0.08 239 74.8 ± 3.0 86 24 7.45 ±0.04 199 61.7 ± 6.2 71 ^(a)Potency is expressed as a pD2 value, which isthe negative logarithm of the concentration required to produce 50% ofthe maximum response each compound (EC50). Relative potency is expressedas a percentage the pD2 value of each compound against that of morphine.Maximum inhibition (%), which is elicited by the compound when theresponse reaches a plateau, was calculated by regarding the twitchcontraction as 100%. Relative inhibitory activity, which means intrinsicactivity on opioid receptors, is expressed as a percentage of themaximum inhibition by each compound against that by morphine. Each valuerepresents a mean (SEM) of five or six animals. The asterisk (*) donatesvalues that were significantly different from the morphine group byStudent's t-test (**, <0.01). Compounds 13a, 15, 17-21, and 25 did notshow significant inhibition at 1 μM.

Among the EG-bridged derivatives (3, 11, 12a, 13a, 14a and 15) and the7-hydroxyindolenine derivatives (22-25) C10-fluorinated derivatives (11,22) showed the highest potency. Derivatives having a fluoro group at C10showed the highest potency amongst the C10-halogenated derivatives.Although not desiring to be bound to any theory, these results suggestthat the dimension or electronegativity of the functional group at theC10 position can be useful to elicit opioid agonistic effect. Compound22 in particular showed potent agonistic effect, but its potency wasnearly equal to that of 7-hydroxymitragynine (1).

The results of these experimental studies show a new method to mask the2, 3-7n bond of indole alkaloids and the conversion of the protectedcompounds, i.e., 2,3-ethylene glycol adducts, back to the startingindoles. This procedure was utilized in these studies for themodification of the benzene ring of the indoline derivative and wasapplied to the preparation of potent opioid receptor agonists with theCorynanthe skeleton, in which the C10-halogenated indole alkaloidcompound exhibited more potent opioid agonistic effect than morphine inin vitro experiments.

Example 2 Indole Alkaloid Compounds of Formula Ia Synthesis of IndoleAlkaloid Compounds of Formula Ia

Exemplary indole alkaloid derivative compounds of formulae Ia wereprepared by reacting 7-hydroxymitragynine and separately7-hydroxy-10-halomitragynine with NaBH₄, in MeOH, to yield compounds 26and 27, such as shown in general synthesis Scheme 4.

Additional details on the experimental procedures used for thepreparation of compounds 26 and 27, and ¹H and ¹³C NMR spectral data forthese compounds are provided below.

Preparation of Compound 26. To a stirred solution of7-hydroxymitragynine (37.5 mg, 0.091 mmol) in dry MeOH (1.1 mL) wasadded NaBH₄ (4.3 mg, 0.11 mmol) at 0° C. under argon atmosphere. After30 min, H₂O was added to the reaction mixture. The mixture wasconcentrated under reduced pressure and poured into saturated aqueousNaHCO₃ solution, and the whole mixture was extracted with 5% MeOH/CHCl₃three times. The combined extract was washed with brine, dried overMgSO₄, and evaporated to give a residue that was purified by silica gelcolumn chromatography (ethyl acetate/n-hexane=35:65) and thencrystallized from ethyl acetate to give 26.3 mg of compound 26 (MGM-15)(y. 70%).

Compound 26; m.p.: 219-223° C. (ethyl acetate). UV (MeOH) λ_(max) nm(log ε): 288 (3.13), 277 (3.06), 239 (4.16), 232 (4.14), 214 (4.50). IR(KBr) ν_(max) cm⁻¹: 3341, 2954, 1698, 1614, 1467, 1283. ¹H-NMR (500 MHz,CDCl₃) δ ppm: 7.40 (1H, s, H-17), 7.00 (1H, dd, J=8.1, 8.1 Hz, H-11),6.35 (1H, d, J=7.9 Hz, H-12), 6.31 (1H, d, J=8.2 Hz, H-10), 3.87 (1H,br.s, N, —H), 3.83 (3H, s, 9-OCH₃), 3.80 (3H, s, 17-OCH₃), 3.70 (3H, s,22-OCH₃), 3.47 (1H, br.s, H-2), 2.95 (2H, m, H-15 and H-21), 2.94 (1H,s, 7-OH), 2.53 (2H, m, H-5 and H-14), 2.25 (1H, ddd, J=12.3, 12.3, 2.3Hz, H-5), 2.16 (2H, m, H-3 and H-21), 2.03 (1H, d, J=14.3 Hz, H-6), 1.92(1H, ddd, J=13.6, 13.6, 4.3 Hz, H-6), 1.79 (1H, m, H-19), 1.58 (1H,br.d, J=11.3 Hz, H-20), 1.39 (1H, d, J=12.8 Hz, H-14), 1.23 (1H, m,H-19), 0.85 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δppm: 169.1 (C-22), 160.3 (C-17), 155.9 (C-9), 149.9 (C-13), 129.3(C-11), 121.3 (C-8), 111.8 (C-16), 105.2 (C-12), 101.9 (C-10), 77.1(C-7), 69.8 (C-2), 61.6 (17-OCH₃), 61.6 (C-3), 58.5 (C-21), 55.1(9-OCH₃), 51.2 (22-OCH₃), 50.7 (C-5), 40.7 (C-20), 40.2 (C-15), 35.2(C-6), 28.5 (C-14), 19.1 (C-19), 13.1 (C-18). EI-MS (%) m/z: 416 (M⁺,61), 400 (96), 399 (100), 398 (97), 397 (78), 383 (41), 256 (64), 214(84). Anal. Calcd for C₂₃H₃₂O₅N₂: C, 66.32; H, 7.74; N, 6.73. Found: C,66.08; H, 7.77; N, 6.71. CD (c=0.29 mM, MeOH, 24° C.), Δε (λnm): 0(318), (+1.4 (293), 0 (257), −1.0 (248), 0 (233), +0.2 (231), 0 (227),−7.7 (215), +0.1 (208).

Preparation of Compound 27. To a stirred solution of10-fluoro7-hydroxymitragynine (23.7 mg, 0.055 mmol) in dry MeOH (0.5 mL)was added NaBH₄ (2.1 mg, 0.056 mmol) at 0° C. under argon atmosphere andthe reaction mixture was stirred for 30 min. After adding H₂O, thereaction mixture was poured into saturated aqueous NaHCO₃ solution. Thewhole mixture was extracted with 5% MeOH/CHCl₃ three times. The combinedextract was washed with brine, dried over MgSO₄, and evaporated to givea residue that was purified by amino-silica get column chromatography(ethyl acetate/n-hexane=1:1) to give 22.6 mg of compound 27 (MGM-16) (y.95%).

Compound 27; UV (MeOH) λ_(max) nm (log ε): 297 (3.13), 276 (3.10), 240(4.22), 226 (4.11), 205 (4.60). IR (KBr) ν_(max) cm⁻¹: 3364, 2947, 1701,1626, 1490, 1284, 1240. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.41 (1H, s,H-17), 6.76 (1H, dd, J=12.8, 8.2 Hz, H-11), 6.28 (1H, dd, J=8.5, 3.1 Hz,H-12), 4.01 (3H, d, J=2.7 Hz, 9-OCH₃), 3.79 (3H, s, 17-OCH₃), 3.69 (3H,s, 22-OCH₃), 3.46 (1H, d, J=3.1 Hz, H-2), 2.95 (2H, m, H-15 and H-21),2.77 (1H, br.s, 7-OH), 2.52 (2H, m, H-5 and H-14), 2.25 (1H, m, H-5),2.15 (2H, m, H-3 and H-21), 1.95 (2H, m, H-6), 1.76 (1H, m, H-19), 1.58(1H, br.d, J=11.3 Hz, H-20), 1.37 (1H, d, J=12.8 Hz, H-14), 1.23 (1H, m,H-19), 0.84 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δppm: 169.0 (C-22), 160.3 (C-17), 148.4 (d, J=237.4 Hz, C-10), 145.1(C-13), 143.2 (d, J=11.9 Hz, C-9), 126.8 (C-8), 116.4 (d, J=21.5 Hz,C-11), 111.7 (C-16), 105.8 (d, J=6.9 Hz, C-12), 77.4 (C-7), 70.0 (C-2),61.6 (17-OCH₃), 61.4 (C-3), 61.2 (d, J=7.8 Hz, 9-OCH₃), 58.4 (C-21),51.2 (22-OCH₃), 50.5 (C-5), 40.7 (C-20), 40.1 (C-15), 35.2 (C-6), 28.4(C-14), 19.0 (C-19), 13.1 (C-18). CD (c=0.26 mM, MeOH, 24° C.), Δε(λnm): 0 (337), +2.7 (297), 0 (255), −4.2 (236), −3.8 (227), −8.4 (214),+0.1 (207). FAB-MS (NBA) m/z: 435 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd.for C₂₃H₃₂O₅N₂F: 435.2295, found: 435.2301.

Pharmacological Evaluation Methods

Animals. Male ddY-strain mice (Japan SLC, Hamamatsu, Japan) weighing25-33 g and male Hartley-strain guinea-pigs (Japan SLC) weighing 320-550g were used.

Drugs. For guinea-pig isolated ileum test, compound 1 and compound 11were first dissolved in 100% dimethylsulfoxide to yield a 5 mM solution,and then subsequently diluted with distilled water. For mouse tail-flicktest and gastrointestinal transit test, compound 1 and compound 11 weredissolved in 25 mM phosphate buffer (pH 5.3-5.5).

Electrical stimulation of guinea-pig ileum. These tests were alsoconducted on compounds 26 and 27, in addition to the compoundspreviously reported in Table A, and the combined results are set forthin Table 1. The guinea-pig ileum was dissected and placed inKrebs-Henseleit solution (in mM: NaCl, 112.08; KCl, 5.90; CaCl₂, 1.97;MgCl₂, 1.18; NaH₂PO₄, 1.22; NaHCO₃, 25.00, and glucose, 11.49). Theileum was placed under 1 g tension in a 5 ml organ bath containing thenutrient solution. The bath was maintained at 37° C. and continuouslybubbled with a mixture of 95% O₂ and 5% CO₂. Tissues were stimulated bya platinum needle-ring (the ring was placed 20 mm above the base of a 5mm long needle) electrode. After equilibration, the ileum wastransmurally stimulated with monophasic pulses (0.2 Hz and 0.1 msduration) by a stimulator (SEN-7203, Nihon Kohden, Tokyo, Japan).Contractions were isotonically recorded by using a displacementtransducer (NEC Type 45347, San-ei Instruments Ltd., Tokyo, Japan). Theeffects of drug treatments on the twitch contractions evoked bytransmural stimulation elicited through the ring electrodes wereexamined. The height of the twitch response to transmural stimulationwas measured before and after the drug challenge. The responses wereexpressed as % inhibition of the twitch response to the transmuralstimulation before the drug challenge.

Tail-flick test. Mice respond to a focused heat stimulus by flicking ormoving their tail from the path of the stimulus, thereby exposing aphotocell located in the tail-flick analgesia meter (Ugo BasileTail-flick Unit 7360, Ugo Basile, Comerio, Italy) immediately below thetail. The reaction time is automatically recorded. Prior to treatmentwith drugs, vehicle, or saline, the nociceptive threshold was measuredthree times, and the mean of the reaction time was used as the pre-druglatency for each mouse. A cut-off time of 10 s was used to preventtissue damage. Analgesia was quantified using the percentage of maximumpossible effect (% MPE) and calculated as: % MPE=[(test latency−pre-druglatency)/(cut-off time−pre-drug latency)]×100.

Gastrointestinal transit. Mice were fasted, with water available adlibitum, for 18 h before the experiments. Fifteen minutes after s.c.administration of compound 1, compound 11, morphine, vehicle, or saline,a charcoal meal (an aqueous suspension of 10% charcoal and 5% gumarabic) was orally administered at a volume of 0.25 ml. Fifteen minutesafter oral administration of compound 1, compound 11 or vehicle, and 30min after oral administration of morphine or distilled water, a charcoalmeal was orally administered. Thirty minutes after administration of thecharcoal meal, the animal was sacrificed by cervical dislocation, andthe small intestine from the pylorus to the ileocecum was carefullyremoved. Both the length of the small intestine from the pylorus to theileocecum and the farthest distance to which the charcoal meal hadtraveled were measured. For each animal, the gastrointestinal transit(GIT) was calculated as the percentage of distance traveled by thecharcoal meal relative to the total length of the small intestine. Theinhibition of gastrointestinal transit (%) was calculated as: Inhibitionof gastrointestinal transit (%)=[(saline or vehicle GIT−drugGIT)/(saline or vehicle GIT)]×100.

Statistical analysis. The data are expressed as the mean±S.E.M.Statistical analyses were performed with two-tailed Student's t-test forcomparison of two groups, and by a one-way analysis of variance followedby a Bonferroni multiple comparison test for comparison of more than twogroups. A P value <0.05 was considered statistically significant. ED₅₀values and 95% confidence limits were determined using theLitchfield-Wilcoxon method (1949).

Results and Discussion. Analgesic effects of compound 1, compound 11,compound 26, compound 27 and morphine were investigated in acute thermalpain tests in mice. The oral administration of compound 1, compound 11,compound 26 and compound 27 exhibited potent and dose-related analgesiceffects in the tail-flick test (FIG. 1). The ED₅₀ values (95% confidencelimits) for the analgesic effects of the oral administration of compound1, compound 11, compound 26 and compound 27 were 4.43 mg/kg (1.57-6.93mg/kg), 2.84 mg/kg (1.60-5.05 mg/kg), 1.26 mg/kg (0.84-1.88 mg/kg) and0.26 mg/kg (0.17-0.42 mg/kg), respectively, in the tail-flick test.Compound 1, compound 11, compound 26 and compound 27 elicit 14, 22, 50and 240 times, respectively, more potent analgesic effects than morphinein the mouse tail-flick test (Table 2). It is hypothesized thatmitragynine-related compounds have a favorable structure for inducingthe orally-active analgesic effect.

Site of action in analgesic effects of compounds. The in vivo analgesictest revealed that compound 11, compound 26 and compound 27 showeddose-dependent and strong analgesic effects when orally administered tomice. In order to determine the opioid receptor type selectivity ofanalgesia induced by compound 11, compound 26 and compound 27, mice werepretreated with selective opioid receptor type antagonists in mousetail-flick test. Doses of compound 11, compound 26 and compound 27 wereused and produced a response of 80-90% to detect the effects of theantagonists easily. The analgesic effects of subcutaneously-administeredcompound 11, compound 26 and compound 27 were completely blocked by thenon-selective opioid antagonist naloxone (data not shown).

This result suggests the involvement of opioid receptors in theiranalgesic effects. The effect of compound 11 was markedly inhibited bythe μ-opioid receptor selective antagonist β-funaltrexamine and by theκ-opioid receptor selective antagonist norbinaltorphimine (FIG. 2A). Theselective δ-antagonist naltrindole was ineffective on compound11-induced effect. Taken together, these results indicated that thepotent analgesic effect of compound 11 resulted from its combinedagonistic action on both μ- and κ-opioid receptors.

On the other hand, the effects of compound 26 and compound 27 weremarkedly inhibited by the μ-opioid receptor selective antagonistβ-funaltrexamine, but were moderately inhibited by the δ-opioid receptorselective antagonist naltrindole (FIGS. 2B, C). The selectiveκ-antagonist nor-binaltorphimine had no effect on their effects.Consequently, these results showed that the potent analgesic effects ofcompound 26 and compound 27 are attributed to its combined agonisticaction on both μ- and δ-opioid receptors.

Weaker constipating effects of compounds than that of morphine.Constipation is a major problem during chronic administration of opioidanalgesics such as morphine. The dose required for its analgesic effectis much higher than that required for its constipating effect.Consequently, constipation is not a clinically negligible issue whenmorphine is used for the analgesic drug. Then, the inhibition ofgastrointestinal transit was studied to evaluate the constipating effectof compound 11 in comparison to morphine and compound 1. The effect ofcompound 11 on the passage of a charcoal meal was examined after itsoral administration in mice. The oral administration of the compound 1,compound 11 and morphine dose-dependently inhibited gastrointestinaltransit (FIG. 3). The constipating ED₅₀ values (95% confidence limits)for compound 1, compound 11 and morphine were 7.50 mg/kg (3.95-14.2),11.1 mg/kg (5.96-20.7) and 11.7 mg/kg (5.56-24.6), respectively (Table2).

The constipating effect of compound 11 was equipotent with morphine andcompound 1. The constipating ED₅₀ value of compound 11 was 4 timeslarger than that of its analgesic ED₅₀ (Table 2). In the case ofmorphine, its constipating ED₅₀ value was 5 times smaller than itsanalgesic ED₅₀ value. These results suggest that compound 11 inducesconstipation much less potently than morphine at equi-analgesic dose.

The oral administration of compound 11, compound 26 and compound 27induced potent analgesic effects in mice. It was surprising that theanalgesic effect of compound 27 is 240 fold more potent than that ofmorphine in their oral administration. The effect of compound 11 wasmeditated by its agonistic action on both μ- and κ-opioid receptors,while the effect of compound 26 and compound 27 were meditated by theiragonistic action on both μ- and δ-opioid receptors. In thegastrointestinal transit study, compound 11 inhibited gastrointestinaltransit, but its constipating effect was much weaker than that ofmorphine at equi-analgesic doses.

The dual acting μ- and κ-opioid agonist, compound 11, and the dualacting μ- and δ-opioid agonists, compound 26 and compound 27, arepromising novel analgesics that have more potent analgesic and weakeradverse effects than morphine in clinical management of pain.

TABLE 1 Opioid effects of mitragynine derivatives on twitch contractioninduced by electrical stimulation in guinea-pig isolated ileum pD₂ valueRelative Maximum Inhibitory Compound (−log M) Potency (%) inhibition (%)activity (%) Morphine 7.15 ± 0.05 100 87.2 ± 1.8 100 Ethylene GlycolBridged Derivatives  3 7.70 ± 0.10 354  35.0 ± 11.0 40 11 8.40 ± 0.021778 83.4 ± 3.2 96 12a 7.61 ± 0.17 288 48.1 ± 9.3 55 14a 7.88 ± 0.18 53765.0 ± 4.3 75 Mitragynine Derivatives  2 6.50 ± 0.06 22 72.0 ± 5.0 837-Hydroxyindolenine Derivatives  1 7.78 ± 0.10 426 90.8 ± 3.4 104 227.87 ± 0.04 524 82.5 ± 1.8 95 23 7.53 ± 0.08 239 74.8 ± 3.0 86 24 7.45 ±0.04 199 61.7 ± 6.2 71 26 8.26 ± 0.05 1259 78.2 ± 3.4 90 27 8.81 ± 0.094571 89.0 ± 2.3 102 Potency is expressed as a pD₂ value, which is thenegative logarithm of the concentration required to produce 50% of themaximum response each compound (EC₅₀). Relative potency is expressed asa percentage the pD₂ value of each compound against that of morphine.Maximum inhibition (%), which is elicited by the compound when theresponse reaches a plateau, was calculated by regarding the twitchcontraction as 100%. Relative inhibitory activity, which means intrinsicactivity on opioid receptors, is expressed as a percentage of themaximum inhibition by each compound against that by morphine. Each valuerepresents mean ± S.E.M. of data obtained from five or six animals.Compounds 13a, 15, 17-21, and 25 did not show significant inhibition at1 μM.

TABLE 2 Analgesic effects (tail-flick) and inhibitory effects ongastrointestinal transit (GIT) produced by orally administration ofmorphine, compound 1, compound 11, compound 26 and compound 27 in miceCompound 1 Compound 11 Compound 26 Compound 27 Morphine Tail-flick 4.43(1.57-6.93) 2.84 (1.60-5.05) 1.26 (0.84-1.88) 0.263 (0.165-0.420) 63.0(37.2-106.8) GIT 7.50 (3.95-14.20) 11.1 (6.0-20.7) N.D. N.D. 11.7(5.6-24.6) ED₅₀ represents the median effective dose (mg/kg) (95%confidence limits). N.D.: Not determined.

The disclosures of each patent, patent application and publication citedor described in this document are hereby incorporated herein byreference, in their entirety.

Applicants specifically incorporate the entire contents of all citedreferences in this disclosure. Further, when an amount, concentration,or other value or parameter is given as either a range, preferred range,or a list of upper preferable values and lower preferable values, thisis to be understood as specifically disclosing all ranges formed fromany pair of any upper range limit or preferred value and any lower rangelimit or preferred value, regardless of whether ranges are separatelydisclosed. Where a range of numerical values is recited herein, unlessotherwise stated, the range is intended to include the endpointsthereof, and all integers and fractions within the range. It is notintended that the scope of the invention be limited to the specificvalues recited when defining a range.

Other embodiments of the present teachings will be apparent to thoseskilled in the art from consideration of the present specification andpractice of the present teachings disclosed herein. It is intended thatthe specification and examples be considered as exemplary only and notbe limiting of the claims. All cited references, patents, and patentapplications are incorporated in their entireties herein by reference.

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Supporting Information For Examples Example 1

General

UV: recorded in MeOH on a JASCO V-560 instrument. IR: recorded on aJASCO FT/1R-230 spectrophotometer. ¹H and ¹³C NMR: recorded on a JEOLJNM A400, JNM A-500, JNM ECP-400 or JNM ECP-600 spectrometer, J valuesare given in Hz. EI-MS: direct probe insertion at 70 eV recorded on aJEOL JMS GC-mate spectrometer. FAB-MS: recorded on a JEOL JMS-AX500 orJMS-HX110 mass spectrometer. Optical rotation: measured with a JASCOP-1020 polarimeter. CD: measured with a JASCO J-720WI. Elementalanalysis: performed on a Perkin-Elmer 240, 2408 Elemental Analyzer.Melting point: measured with a Yanagimoto Micro Melting Point Apparatus1631A. TLC: precoated Kieselgel 60 F₂₅₄ plates (Merck, 0.25 mm thick).Column chromatography: Kieselgel 60 [Merck, 70-230 mesh (for openchromatography) and 230-400 mesh (for flash chromatography)],Chromatorex NH [Fuji Silysia Chemical, 100-200 mesh (for amino-silicagel column chromatography)]. Medium pressure liquid chromatography(MPLC): C. I. G. prepacked column CPS-HS-221-05 (Kusano Kagakukikai,SiO₂).

Preparation of Ethylene Glycol-Bridged Compound 3 from Mitragynine (2)

To a solution of mitragynine (2, 102 mg, 0.26 mmol) in dry MeCN (2.0 mL)were added dry ethylene glycol (2.0 mL) and PIFA (113.5 mg, 0.26 mmol)at 0° C. and the mixture was stirred for 1 h at 0° C. under argonatmosphere. After adding chilled aqueous NaHCO₃ solution, the mixturewas extracted three times with chloroform. The combined extract waswashed with brine, dried over MgSO₄, and evaporated. The crude productwas chromatographed on amino-silica gel (ethyl acetate/n-hexane=30:70)to give 120.4 mg (quantitative) of 3 as a colorless amorphous powder, aportion of which was recrystallized from acetone to give pale yellowishprisms (mp. 169-172° C.). Ethylene glycol-bridged Compound 3; UV (MeOH)λ_(max) nm (log ε): 285 (3.35), 277 (3.32), 238 (4.26), 229 (4.22), 213(4.64). IR (ATR) ν_(max) cm⁻¹: 3255, 2929, 1694, 1646, 1605, 1458, 1255.¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.41 (1H, s, H-17), 7.08 (1H, dd, J=8.1,8.1 Hz, H-11), 6.39 (1H, d, J=8.2 Hz, H-40), 6.37 (1H, d, J=7.9 Hz,H-12), 4.25 (1H, brs, N_(a)—H), 3.93 (1H, ddd, J=11.8, 11.8, 2.7 Hz,H-2′), 3.85 (3H, s, 9-OCH₃), 3.84 (1H, m, H-1′, overlapped with 9-OCH₃),3.79 (3H, s, 17-OCH₃), 3.70 (3H, s, 22-OCH₃), 3.67 (1H, dd, J=11.6, 2.4Hz, H-1′), 3.42 (1H, dd, J=11.7, 2.3 Hz, H-2′), 2.98 (1H, dd, J=11.4,2.0 Hz, H-21), 2.92 (1H, ddd, J=13.4, 3.6, 3.6 Hz, H-15), 2.48 (1H, m,H-5), 2.46 (1H, m, H-3), 2.38-2.29 (2H, m, H-5 and H-14), 2.26 (1H, dd,J=11.3, 2.7 Hz, H-21), 2.15 (1H, ddd, J=14.6, 2.4, 2.4 Hz, H-6), 1.85(1H, m, H-6), 1.80 (1H, m, H-14), 1.73 (1H, m, H-19), 1.56 (1H, br. d,J=11.3 Hz, H-20), 1.25 (1H, m, H-19), 0.84 (3H, dd, J=7.3, 7.3 Hz,R₃-18). ¹³C-NMR (100 MHz, CDCl₃) δ ppm: 169.1 (C-22), 160.4 (C-17),156.9 (C-9), 149.0 (C-13), 129.9 (C-11), 115.8 (C-8), 111.7 (C-16),105.3 (C-12), 102.8 (C-10), 90.8 (C-2), 81.3 (C-7), 62.3 (C-1′), 61.6(17-OCH₃), 61.2 (C-2′), 60.8 (C-3), 58.5 (C-21), 55.3 (9-OCH₃), 51.3(22-OCH₃), 50.2 (C-5), 40.5 (C-20), 40.1 (C-15), 35.4 (C-6), 24.2(C-14), 19.2 (C-19), 13.1 (C-18). CD (c=0.21 mM, MeOH, 24° C.), Δε(λnm): 0 (303), −2.4 (286), 0 (263), +0.7 (256), 0 (250), −1.0 (233),−30.2 (211), −9.4 (200). FAB-MS (NBA) m/z: 459 [M+H]⁺. HR-FAB-MS(NBA/PEG): calcd. for C₂₅H₃₅N₂O₆: 459.2495, found: 459.2515. Anal. Calcdfor C₂₅H₃₄N₂O₆.H₂O: C, 63.00; H, 7.61; N, 5.88. Found: C, 63.26; H,7.75; N, 5.73. FIG. 4 a is the 1H-NMR for Ethylene Glycol-BridgedCompound 3 and FIG. 4 b is the 13C-NMR for Ethylene Glycol-BridgeCompound 3.

Conversion of 3 to Mitragynine (2)

To a stirred solution of 3 (24.6 mg, 0.05 mmol) in dry AcOH (0.6 mL) wasadded NaCNBH₃ (19.2 mg, 0.29 mmol) at room temperature and the mixturewas stirred for 2 h at room temperature under argon atmosphere. MeOH (20μL) was added to the reaction mixture, which was then heated underreflux at 90° C. for 4 h. The reaction mixture was poured into coldaqueous NH₄OH solution, and this was extracted three times withchloroform. The combined extract was washed with brine, dried overMgSO₄, and evaporated to give a residue that was purified by silica gelcolumn chromatography (ethyl acetate/n-hexane=50:50) to give 21.7 mg(quantitative) of mitragynine (2), which was completely identical withthe authentic sample.

Preparation of Compound 4

To a stirred solution of 2,3-dimethylindole (74.3 mg, 0.51 mmol) in dryMeCN (2.6 mL) and dry ethylene glycol (2.6 mL) were successively addedNH₄Cl (132.2 mg, 2.45 mmol) and PIFA (251.5 mg, 0.61 mmol) at 0° C. andthe mixture was stirred for 1 h at 0° C. under argon atmosphere. Afteradding chilled aqueous NaHCO₃ solution, the mixture was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated. The crude product was purified by pre-packedsilica gel column chromatography (ethyl acetate/n-hexane=30:70) to give83.1 mg (79%) of 4 as a yellowish amorphous powder, a portion of whichwas recrystallized from n-hexane to give yellowish prisms (mp. 79-81°C., cap.). Compound 4; UV (MeOH) λ_(max) nm: 290, 269, 241, 233, 220,218, 104. IR (KBr) ν_(max) cm⁻¹: 3329, 2979, 2960, 2864, 1612, 1468.¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.21 (1H, d, J=7.3 Hz, H-5), 7.15 (1H,ddd, J=7.7, 7.7, 1.3 Hz, H-7), 6.86 (1H, ddd, J=7.4, 7.4, 0.9 Hz, H-6),6.71 (1H, d, J=7.9 Hz, H-8), 4.11 (1H, br. s, N_(a)—H), 3.72 (2H, m,H-1′ and H-2′), 3.65 (1H, m, H-2′), 3.58 (1H, m, H-1′), 1.56 (3H, s,2-CH₃), 1.38 (3H, s, 3-CH₃). ¹³C-NMR (100 MHz, CDCl₃) δ ppm: 147.6(C-9), 131.1 (C4), 129.0 (C-7), 123.1 (C-5), 119.8 (C-6), 110.5 (C-8),93.2 (C-2), 81.5 (C-3), 61.3 (C-2′), 61.1 (C-1′), 23.3 (2-CH₃), 18.0(3-CH₃). [α]_(D) ²⁵ 0 (c 1.0, CHCl₃). FAB-MS (NBA) m/z: 206 [M+H]⁺.HR-FAB-MS (NBA/PEG): calcd. for C₁₂H₁₆NO₂: 206.1181, found: 206.1180.FIG. 5 a is the 1H-NMR for Compound 4 and FIG. 5 b is the 13C-NMR forCompound 4.

Preparation of Compound 5

To a stirred solution of tetrahydrocarbazole (110.9 mg, 0.65 mmol) indry MeCN (3.3 mL) and dry ethylene glycol (3.3 mL) were successivelyadded NH₄Cl (167.8 mg, 2.60 mmol) and PIFA (335.6 mg, 0.78 mmol) at 0°C. and the mixture was stirred for 30 min at 0° C. under argonatmosphere. After adding chilled aqueous NaHCO₃ solution, the mixturewas extracted three times with chloroform. The combined extract waswashed with brine, dried over MgSO₄, and evaporated. The crude productwas chromatographed on silica gel (ethyl acetate/n-hexane=0:100-50:50)to give 129.5 mg (87%) of 5 as a colorless amorphous powder, a portionof which was recrystallized from ethyl acetate to give colorless prisms(mp. 156-157° C., cap.). Compound 5; UV (MeOH) λ_(max) nm: 295, 275,247, 223, 205. IR (KBr) ν_(max) cm⁻¹: 3311, 3035, 2958, 2933, 2864,1616. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.18 (1H, d, J=7.3 Hz, H-9), 7.14(1H, ddd, J=7.6, 7.6, 1.2 Hz, H-11), 6.85 (1H, dd, J=7.4, 7.4 Hz, H-10),6.71 (1H, d, J=7.6 Hz, H-12), 4.01 (1H, br. s, N_(a)—H), 3.81 (2H,overlapped, H-1′ and H-2′), 3.65 (2H, overlapped, H-1′ and H-2′), 2.12(1H, ddd, J=10.4, 10.4, 2.7), 2.00 (1H, m), 1.70 (2H, m), 1.52 (3H, m),1.42 (1H, m). ¹³C-NMR (100 MHz, CDCl₃) δ ppm: 147.4 (C-13), 131.6 (C-8),128.8 (C-11), 122.4 (C-9), 119.7 (C-10), 111.1 (C-12), 92.3 (C-2), 80.6(C-7), 61.4 (C-1′), 61.0 (C-2′), 36.4, 29.4, 22.9, 20.6. [α]_(D) ²⁵+1 (c1.0, CHCl₃). FAB-MS (NBA) m/z: 231 [M]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₁₄H₁₇NO₂: 231.1259, found: 231.1269. FIG. 6 a is the 1H-NMR forCompound 5 and FIG. 6 b is the 13C-NMR for Compound 5.

Preparation of Compound 6

To a stirred solution of indoloquinolizidine (70.4 mg, 0.31 mmol) in dryMeCN (1.6 mL) and dry ethylene glycol (1.6 mL) were successively addedNH₄Cl (83.2 mg, 1.55 mmol) and PIFA (157.8 mg, 0.37 mmol) at roomtemperature and the mixture was stirred for 2 h at 50° C. under argonatmosphere, After adding chilled aqueous NaHCO₃ solution, the mixturewas extracted three times with chloroform. The combined extract waswashed with brine, dried over MgSO₄, and evaporated. The crude productwas purified by pre-packed silica gel column chromatography (5% MeOH inCHCl₃) to give 86.0 mg (97%) of 6 as a colorless amorphous powder, aportion of which was recrystallized from acetone and n-hexane to givecolorless prisms (mp. 142-143° C., cap.). Compound 6; UV (MeOH) λ_(max)nm: 290, 262, 238, 221, 206. IR (ATR) ν_(max) cm⁻¹: 3313, 3033, 2964,2939, 2924, 2910, 1614. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.18 (1H, dd,J=7.2, 0.6 Hz, H-9), 7.14 (1H, ddd, J=7.6, 7.6, 1.3 Hz, H-11), 6.86 (1H,ddd, J=7.4, 7.4, 0.9 Hz, H-10), 6.71 (1H, d, J=7.7 Hz, H-12), 4.20 (1H,br. s, N_(a)—H), 3.90 (2H, overlapped, H-1′ and H-2′), 3.61 (1H, m, H-1′or H-2′), 3.44 (1H, m, H-1′ or H-2′), 2.99 (1H, d, J=11.0 Hz), 2.57 (1H,m), 2.50 (1H, dd, J=10.9, 2.1 Hz, H-3), 2.41 (1H, ddd, J=12.3, 12.3, 2.5Hz), 2.16 (1H, ddd, J=11.1, 11.1, 4.3 Hz), 2.05 (1H, d, J=10.3 Hz), 1.91(2H, m), 1.80 (1H, ddd, J=13.8, 13.8, 4.5 Hz), 1.68 (2H, m), 1.43 (1H,m), 1.33 (1H, m). ¹³C-NMR (100 MHz, CDCl₃) δ ppm: 146.8 (C-13), 130.8(C-8), 128.6 (C-11), 122.0 (C-9), 119.9 (C-10), 111.8 (C-12), 91.1(C-2), 80.4 (C-7), 62.0 and 61.2 (C-1′ and C-2′), 59.5, 56.5, 50.6,37.4, 25.5, 24.8, 24.6. [α]_(D) ²⁴+1 (c 0.8, CHCl₃). FAB-MS (NBA) m/z:287 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₁₇H₂₃N₂O₂: 287.1760, found:287.1762. FIG. 7 a is the ¹H-NMR for Compound 6 and FIG. 7 b is the¹³C-NMR for Compound 6.

Preparation of Compound 7

To a stirred solution of dihydrocorynantheol (229 mg, 0.77 mmol) in dryMeCN (4.0 mL) and dry ethylene glycol (4.0 mL) was added NH₄Cl (55.4 mg,1.04 mmol), and PIFA was added portionwise to the stirred mixture atroom temperature under argon atmosphere in the following manner: 0 min,157.6 mg (0.37 mmol); 1 h, 175.7 mg (0.41 mmol); 4 h, 37.5 mg (0.08mmol). The reaction mixture was stirred at the same temperature forfurther 3 h under argon atmosphere. After adding chilled aqueous NaHCO₃solution, the mixture was extracted three times with chloroform. Thecombined extract was washed with brine, dried over MgSO₄, andevaporated. The crude product was chromatographed on amino-silica gel(ethyl acetate/chloroform=30:70) to give 238.7 mg (87%) of 7 as acolorless amorphous powder, a portion of which was recrystallized fromethyl acetate and n-hexane to give colorless prisms (mp. 108-111° C.cap.). Compound 7; UV (MeOH) λ_(max) nm: 292, 264, 239, 221, 205. IR(ATR) ν_(max) cm⁻¹: 3313, 2958, 2866, 1614, 1466. ¹H-NMR (500 MHz,CDCl₃) δ ppm: 7.17 (1H, d, J=7.0 Hz, H-9), 7.13 (1H, ddd, J=7.6, 7.6,1.2 Hz, H-11), 6.86 (1H, ddd, J=7.3, 7.3, 0.9 Hz, H-10), 6.71 (1H, d,J=7.6 Hz, H-12), 4.23 (1H, br. s, N_(a)—H), 3.89 (2H, overlapped, H-1′and H-2′), 3.75 (2H, m, H₂-17), 3.61 (1H, m, H-1′), 3.44 (1H, m, H-2′),3.05 (1H, dd, J=11.3, 3.6 Hz, H-21), 2.60 (1H, dddd, J=11.6, 2.2, 2.2,2.2 Hz, H-5), 2.51 (1H, dd, J=11.1, 1.9 Hz, H-3), 2.39 (1H, ddd, J=12.4,12.4, 2.4 Hz, H-5), 2.15 (1H, ddd, J=12.0, 2.8, 2.8 Hz, H-14), 1.96 (2H,overlapped, H-6 and H-16), 1.88 (1H, dd, J=11.3, 11.3 Hz, H-21), 1.80(1H, ddd, J=13.8, 13.8, 4.4 Hz, H-6), 1.67 (1H, m, H-19), 1.38 (3H,overlapped, H-15, H-16 and H-20), 1.24 (1H, ddd, J=11.6, 11.6, 11.6 Hz,H-14), 1.11 (1H, m, H-19), 0.89 (3H, dd, J=7.5, 7.5 Hz, H₃-18). ¹³C-NMR(100 MHz, CDCl₃) δ ppm: 146.7 (C-13), 130.8 (C-8), 128.7 (C-11), 122.0(C-9), 119.9 (C-10), 111.8 (C-12), 91.1 (C-2), 80.3 (C-7), 62.0 (C-1′),61.3 (C-2′), 61.0 (C-21), 60.5 (C-17), 59.1 (C-3), 50.3 (C-5), 41.5(C-20), 37.4 (C-6), 37.0 (C-15), 35.8 (C-16), 30.5 (C-14), 23.3 (C-19),11.0 (C-18). [α]_(D) ²⁵-72 (c 1.1, CHCl₃). EI-MS m/z (%): 358 (100, M⁺),313 (20), 169 (95), 156 (41). HR-FAB-MS (NBA/PEG): calcd. forC₂₁H₃₁N₂O₃: 359.2335, found: 359.2344.

Preparation of Compound 8

To a stirred solution of corynantheol (302 mg, 1.02 mmol) in dry MeCN(15.0 mL) and dry ethylene glycol (15.0 mL) was added NH₄Cl (107.4 mg,2.01 mmol), and PIFA was added portionwise to the stirred mixture at 0°C. under argon atmosphere in the following manner: 0 min, 139.1 mg (0.32mmol); 10 min, 142.3 mg (0.33 mmol); 20 min, 175.5 mg (0.41 mmol); 30min, 100.5 mg (0.23 mmol). The reaction mixture was stirred at roomtemperature for further 15 h under argon atmosphere. After addingchilled aqueous NaHCO₃ solution, the mixture was extracted three timeswith chloroform. The combined extract was washed with brine, dried overMgSO₄, and evaporated. The crude product was chromatographed onamino-silica gel (ethyl acetate/chloroform=30:70) to give 352.9 mg (97%)of 8 as a yellowish amorphous powder, a portion of which wasrecrystallized from ethyl acetate and n-hexane to give colorless prisms(mp. 105-110° C., cap.). Compound 8; UV (MeOH) λ_(max) nm: 291, 265,239, 222, 205. IR (KBr) ν_(max) cm⁻¹: 3315, 2924, 2866, 2821, 1614,1466. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.18 (1H, d, J=7.3 Hz, H-9), 7.14(1H, ddd, J=7.6, 7.6, 1.3 Hz, H-11), 6.87 (1H, dd, J=7.3, 7.3 Hz, H-10),6.72 (1H, d, J=7.6 Hz, H-12), 5.58 (1H, ddd, J=17.2, 10.2, 8.9 Hz,H-19), 5.12 (1H, dd, J=17.2, 1.7 Hz, H-18), 5.09 (1H, dd, J=10.2, 1.7Hz, H-18), 4.23 (1H, br. s, N_(a)—H), 3.89 (2H, overlapped, H-1′ andH-2′), 3.75 (2H, m, H-17), 3.61 (1H, m, H-1′), 3.44 (1H, m, H-2′), 2.89(1H, dd, J=11.1, 3.7 Hz, H-21), 2.57 (2H, m, overlapped H-3 and H-5),2.40 (1H, ddd, J=12.4, 12.4, 2.4 Hz, H-5), 2.19 (2H, m, overlapped H-14and H-20), 2.07 (1H, dd, J=11.1, 11.1 Hz, H-21), 1.92 (2H, overlapped,H-6 and H-16), 1.79 (1H, ddd, J=13.9, 13.9, 4.6 Hz, H-6), 1.46 (1H, m,H-15), 1.35 (1H, m, H-16), 1.24 (1H, ddd, J=11.4, 11.4, 11.4 Hz, H-14).¹³C-NMR (100 MHz, CDCl₃) δ ppm: 146.7 (C-13), 139.5 (C-19), 130.7 (C-8),128.7 (C-11), 122.1 (C-9), 120.0 (C-10), 116.9 (C-18), 111.8 (C-12),91.0 (C-2), 80.3 (C-7), 61.9 (C-1′), 61.7 (C-21), 61.3 (C-2′), 60.4(C-17), 58.9 (C-3), 50.0 (C-5), 46.8 (C-20), 37.3 (C-6), 36.8 (C-16),36.7 (C-15), 30.1 (C-14). [α]_(D) ²⁴-81 (c 1.0, CHCl₃). EI-MS m/z (%):356 (100, M⁺), 166 (76), 154 (37). HR-FAB-MS (NBA/PEG): calcd. forC₂₁H₂₉N₂O₃: 357.2178, found: 357.2169.

Preparation of Compound 9

To a stirred solution of yohimbine (117.4 mg, 0.33 mmol) in dry MeCN(1.7 mL) and dry ethylene glycol (1.7 mL) were successively added NH₄Cl(69.8 mg, 1.29 mmol) and PIFA (157.0 mg, 0.36 mmol) at room temperatureand the mixture was stirred for 3 h at 40° C. under argon atmosphere.After adding chilled aqueous NaHCO₃ solution, the mixture was extractedthree times with chloroform. The combined extract was washed with brine,dried over MgSO₄, and evaporated. The crude product was chromatographedon silica gel (10% MeOH in CHCl₃) to give 113.2 mg (83%) of 9 as a paleyellowish amorphous powder, a portion of which was recrystallized fromacetone and n-hexane to give colorless prisms (mp. 231-234° C., cap.).Compound 9; UV (MeOH) max nm: 292, 265, 239, 222, 205. IR (KBr) ν_(max)cm⁻¹: 3381, 2924, 1726, 1616, 1468. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.17(1H, d, J=6.8 Hz, H-9), 7.13 (1H, ddd, J=7.6, 7.6, 1.3 Hz, H-11), 6.87(1H, dd, J=7.3, 7.3 Hz, H-10), 6.72 (1H, d, J=7.8 Hz, H-12), 4.19 (1H,br. s, N_(a)—H), 4.18 (1H, m, H-17), 3.87 (2H, overlapped, H-1′ andH-2′), 3.76 (3H, s, 22-OCH₃), 3.60 (1H, d, J=9.3 Hz, H-1′), 3.42 (1H, d,J=9.5 Hz, H-2′), 3.27 (1H, br. s, 17-OH), 2.88 (1H, dd, J=11.1, 3.1 Hz,H-21), 2.61 (1H, d, J=11.5 Hz, H-3), 2.57 (1H, m, H-5), 2.42 (1H, ddd,J=12.3, 12.3, 2.4 Hz, H-5), 2.31 (1H, dd, J=11.1, 1.8 Hz, H-16), 2.02(1H, dd, J=10.9, 10.9 Hz, H-21), 1.99 (1H, m, H-18), 1.90 (3H,overlapped, H-6, H-14 and H-15), 1.77 (1H, ddd, J=13.8, 13.8, 4.4 Hz,H-6), 1.50 (3H, overlapped, H-18, H-19 and H-20), 1.39 (1H, m, H-19),1.24 (1H, ddd, J=12.1, 12.1, 12.1 Hz, H-14). ¹³C-NMR (100 MHz, CDCl₃) δppm: 176.1 (C-22), 146.6 (C-13), 130.7 (C-8), 128.7 (C-11), 122.1 (C-9),120.1 (C-10), 111.8 (C-12), 90.9 (C-2), 80.2 (C-7), 66.6 (C-17), 61.9(C-1′), 61.8 (C-21), 61.3 (C-2′), 59.2 (C-3), 52.3 (C-16), 51.9(22-OCH₃), 50.1 (C-5), 40.3 (C-20), 37.4 (C-6), 36.8 (C-15), 31.1(C-18), 29.3 (C-14), 23.2 (C-19). [α]_(D) ²⁵-44 (c 0.98, CHCl₃). FAB-MS(NBA) m/z: 415 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₃H₃₁N₂O₅:415.2233, found: 415.2202.

Preparation of Compound 10

To a stirred solution of reserpine (100.7 mg, 0.17 mmol) in dry MeCN(2.0 mL) and dry ethylene glycol (2.0 mL) was added IBDA (111.2 mg, 0.31mmol) at 0° C. After 2 h, another portion of IBDA (26.0 mg, 0.07 mmol)was added to the reaction mixture, and this was stirred for 1 h at 0° C.under argon atmosphere. After adding chilled aqueous NaHCO₃ solution,the mixture was extracted three times with chloroform. The combinedextract was washed with brine, dried over MgSO₄, and evaporated. Thecrude product was chromatographed on silica gel (ethylacetate/chloroform=30:70) to give 102.3 mg (93%) of 10 as a yellowishoil, a portion of which was recrystallized from ethyl acetate to givecolorless prisms (mp. 130-132° C., cap.). Compound 10; UV (MeOH) λ_(max)nm: 267, 246, 267. IR (ATR) ν_(max) cm⁻¹: 3355, 2934, 1739, 1717, 1626.¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.32 (2H, s, H-24 and H-28), 7.05 (1H, d,J=8.1 Hz, H-9), 6.43 (1H, dd, J=8.1, 2.2 Hz, H-10), 6.35 (1H, d, J=2.2Hz, H-12), 5.06 (1H, m, H-18), 3.92 (9H, overlapped, 25-OCH₃, 26-OCH₃and 27-OCH₃), 3.90 (1H, s), 3.88 (1H, dd, J=11.7, 1.7 Hz, H-17), 3.83(1H, m, H-2′), 3.81 (3H, s, 16-CO₂CH₃), 3.78 (3H, s, 11-OCH₃), 3.76 (1H,m, H-1′), 3.60 (1H, m, H-1′), 3.58 (1H, m, H-21), 3.54 (3H, s, 17-OCH₃),3.54 (1H, dd, J=9.0, 1.5 Hz), 3.49 (1H, d, J=7.3 Hz), 3.43 (1H, dd,J=11.7, 1.7 Hz, H-2′), 3.10 (1H, ddd, J=13.6, 13.6, 2.2 Hz, H-5), 2.79(1H, dd, J=11.0, 5.1 Hz, H-16), 2.67 (1H, m, H-15), 2.59 (1H, m, H-5),2.16 (3H, overlapped, H-19, H-20 and H-21), 2.00 (1H, m, H-14), 1.95(1H, m, H-19), 1.87 (1H, ddd, J=13.4, 13.4, 4.2 Hz, H-6), 1.80 (1H, dd,J=14.7, 4.5 Hz, H-14), 1.69 (1H, m, H-6). ¹³C-NMR (100 MHz, CDCl₃) δppm: 172.6 (16-CO₂CH₃), 165.4 (C-22), 160.9 (C-11), 152.9 (C-25, C-27),148.4 (C-13), 142.3 (C-26), 125.4 (C-23), 122.9 (C-9), 122.6 (C-8),106.8 (C-24, C-28), 105.7 (C-0), 98.3 (C-12), 89.2 (C-2), 80.9 (C-7),77.9 (C-18), 77.8 (C-17), 61.5 (C-2′), 61.4 (C-1′), 60.9 (26-OCH₃), 60.8(17-OCH₃), 56.2 (25-OCH₃, 27-OCH₃), 55.4 (1-OCH₃), 52.0 (16-CO₂CH₃),51.9 (C-16), 51.3 (C-21), 51.0 (C-3), 48.7 (C-5), 34.0 (C-6), 33.9(C-20), 32.6 (C-15), 30.1 (C-19), 20.1 (C-14). [α]_(D) ²⁴-45 (c 0.99,CHCl₃). FAB-MS (NBA) m/z: 669 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₃₅H₄₅N₂O₁₁: 669.3023, found: 669.3008.

Fluorination of Compound 3

To a solution of 3 (20.6 mg, 0.045 mmol) in dry DCM (0.4 mL) were addedFP-T800 (17.0 mg, 0.054 mmol) and MeNO₂ (47 μL) at 0° C. and the mixturewas stirred at the same temperature under argon atmosphere. After 24 h,FP-T800 (4.8 mg, 0.015 mmol) was added to the reaction mixture, whichwas further stirred for 46.5 h at 0° C. The reaction mixture was pouredinto cold saturated aqueous NaHCO₃ solution, and this was extractedthree times with chloroform. The combined extract was washed with brine,dried over MgSO₄, and concentrated to give a residue that was purifiedby pre-packed amino-silica gel column chromatography (ethylacetate/n-hexane=20:80) to give 11.3 mg (53%) of 11 as a colorlessamorphous powder, together with 2.2 mg (11%) of recovered startingmaterial. Compound 11; UV (MeOH) λ_(max) (log ε) nm: 295 (3.36), 277(3.20), 239 (4.13), 225 (4.06), 207 (4.44). IR (ATR) ν_(max) cm⁻¹: 2926,1697, 1647, 1489, 1242. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.42 (1H, s,H-17), 6.84 (1H, dd, J=12.5, 8.5 Hz, H-11), 6.32 (1H, dd, J=8.4, 3.3 Hz,H-12), 4.16 (1H, br. s, N_(a)—H), 4.00 (3H, d, J=2.4 Hz, 9-OCH₃), 3.91(2H, m, H-1′, H-2′), 3.79 (3H, s, 17-OCH₃), 3.71 (1H, m, H-1′,overlapped with 22-OCH₃), 3.70 (3H, s, 22-OCH₃), 3.44 (1H, d, J=9.5 Hz,H-2′), 3.00 (1H, d, J=11.6 Hz, H-21), 2.91 (1H, ddd, J=13.1, 3.6, 3.6Hz, H-15), 2.50 (1H, ddd, J=11.3, 3.3, 3.3 Hz, H-5), 2.45 (1H, d, J=11.0Hz, H-3), 2.34 (2H, m, H-5 and H-14), 2.26 (1H, dd, J=11.6, 3.1 Hz,H-21), 2.13 (1H, d, J=14.3 Hz, H-6), 1.86 (1H, ddd, J=13.6, 13.6, 4.2Hz, H-6), 1.77 (1H, d, J=12.5 Hz, H-14), 1.72 (1H, m, H-19), 1.57 (1H,br. d, J=1.3 Hz, H-20), 1.24 (1H, m, H-19), 0.84 (3H, dd, J=7.5, 7.5 Hz,H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 169.0 (C-22), 160.4 (C-17),149.5 (d, J=237.8 Hz, C-10), 144.6 (d, J=12.4 Hz, C-9), 144.1 (C-13),121.9 (C-8), 116.7 (d, J=21.5 Hz, C-11), 111.5 (C-16), 106.0 (d, J=7.3Hz, C-12), 91.5 (C-2), 81.5 (C-7), 62.5 (C-1′), 61.6 (17-OCH₃), 61.2(C-2′), 61.1 (d, J=6.9 Hz, 9-OCH₃), 60.8 (C-3), 58.4 (C-21), 51.3(22-OCH₃), 50.2 (C-5), 40.4 (C-20), 40.0 (C-15), 36.0 (C-6), 24.0(C-14), 19.1 (C-19), 13.0 (C-18). ¹⁹F-NMR (564.7 MHz, CDCl₃) δ ppm:−141.5 (d, J=12.9 Hz). CD (c=0.27 mM, MeOH, 24° C.), Δε (λ nm): 0 (319),−0.9 (293), 0 (268), +0.9 (253), 0 (245), −19.0 (208), −14.6 (200).FAB-MS (NBA) m/z: 477 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₂₅H₃₄N₂O₆F: 477.2401, found: 477.2381. FIG. 8 a is the 1H-NMR forCompound 11 and FIG. 8 b is the 13C-NMR for Compound 11.

Chlorination of Compound 3

To a solution of 3 (106 mg, 0.23 mmol) in dry AcOH (2.0 mL) was addedN-chlorosuccinimide (38.7 mg, 0.58 mmol) and the mixture was stirred for12 h at room temperature under argon atmosphere. After adding Na₂S₂O₃,the mixture was poured into cold aqueous NH₄OH solution, and this wasextracted three times with chloroform. The combined extract was washedwith brine, dried over MgSO₄, and evaporated. The crude product waschromatographed on amino-silica get (ethyl acetate/n-hexane=10:90) togive 100.7 mg (88%) of 12a and 13.2 mg (11%) of 12b, as a colorlessamorphous powder, respectively. Compound 12a; UV (MeOH) λ_(max) nm (logε): 299 (3.12), 279 (2.92), 246 (4.18), 227 (3.96), 210 (4.40). IR (ATR)ν_(max) cm⁻¹: 2942, 1701, 1604, 1456, 1285. ¹H-NMR (400 MHz, CDCl₃) δppm: 7.42 (1H, s, H-17), 7.12 (1H, d, J=8.2 Hz, H-11), 6.43 (1H, d,J=8.2 Hz, H-12), 4.30 (1H, br. s, N_(a)—H), 3.95 (1H, m, H-2′), 3.95(3H, s, 9-OCH₃), 3.90 (1H, m, H-1′), 3.79 (3H, s, 17-OCH₃), 3.70 (3H, s,22-OCH₃), 3.70 (1H, m, H-1′), 3.43 (1H, d, J=9.2 Hz, H-2′), 3.00 (1H,dd, J=11.6, 2.1 Hz, H-21), 2.92 (1H, ddd, J=13.2, 3.6, 3.6 Hz, H-15),2.51 (1H, m, H-5), 2.45 (1H, dd, J=11.3, 1.6 Hz, H-3), 2.34 (2H, m, H-5and H-14), 2.26 (1H, dd, J=11.2, 3.3 Hz, H-21), 2.15 (1H, ddd, J=14.3,2.5, 2.5 Hz, H-6), 1.88 (1H, ddd, J=13.8, 13.8, 4.6 Hz, H-6), 1.78 (1H,br. d, J=11.2 Hz, H-14), 1.72 (1H, m, H-19), 1.58 (1H, br. d, J=11.2 Hz,H-20), 1.24 (1H, m, H-19), 0.85 (3H, dd, J=7.4, 7.4 Hz, H₃-18). ¹³C-NMR(100 MHz, CDCl₃) δ ppm: 169.0 (C-22), 160.4 (C-17), 152.9 (C-9), 147.9(C-13), 130.6 (C-11), 123.3 (C-8), 118.0 (C-10), 111.6 (C-16), 108.3(C-12), 91.2 (C-2), 81.5 (C-7), 62.7 (C-1′), 61.7 (17-OCH₃), 61.3(9-OCH₃), 61.1 (C-2′), 60.8 (C-3), 58.5 (C-21), 51.3 (22-OCH₃), 50.2(C-5), 40.4 (C-20), 40.0 (C-15), 36.4 (C-6), 24.0 (C-14), 19.1 (C-19),13.0 (C-18). CD (c=0.25 mM, MeOH, 24° C.), Δε (λ nm): 0 (318), −1.0(292), 0 (267), +0.3 (259), 0 (251), −0.9 (240), −0.8 (236), −24.8(209), −11.4 (200). EI-MS (%) m/z: 494 (M⁺+2, 30), 492 (M⁺, 83) 239(100). HR-FAB-MS (NBA/PEG): calcd. for C₂₅H₃₄N₂O₆ ³⁵Cl: 493.2105, found:493.2104, calcd. for C₂₅H₃₄N₂O₆ ³⁷Cl: 495.2089, found: 495.2061.Compound 12b; UV (MeOH) λ_(max) nm (log ε): 293 (3.24), 276 (3.01), 237(4.13), 231 (4.11), 215 (4.52). IR (ATR) ν_(max) cm⁻¹: 2940, 1698, 1608,1486, 1279, 1238. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.43 (1H, s, H-17),7.07 (1H, dd, J=8.8, 0.5 Hz, H-11), 6.35 (1H, d, J=8.6 Hz, H-10), 4.45(1H, br. s, N_(a)—H), 3.93 (1H, ddd, J=11.7, 11.7, 2.9 Hz, H-2′), 3.83(3H, s, 9-OCH₃), 3.79 (3H, s, 17-OCH₃), 3.78 (1H, m, H-1′, overlappedwith 17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.69 (1H, m, H-1′, overlapped with22-OCH₃), 3.43 (1H, dd, J=11.6, 2.5 Hz, H-2′), 3.00 (1H, d, J=11.5 Hz,H-21), 2.93 (1H, ddd, J=13.0, 3.5, 3.5 Hz, H-15), 2.48 (2H, m, H-3 andH-5), 2.37 (2H, m, H-5 and H-14), 2.26 (1H, dd, J=11.4, 3.3 Hz, H-21),2.15 (1H, ddd, J=14.5, 2.1, 2.1 Hz, H-6), 1.82 (2H, m, H-6 and H-14),1.75 (1H, m, H-19), 1.26 (1H, m, H-19), 0.85 (3H, dd, J=7.3, 7.3 Hz,H₃-18), *H-20: under H₂O. ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 169.2 (C-22),160.5 (C-17), 155.6 (C-9), 145.6 (C-13), 129.2 (C-11), 117.3 (C-8),111.6 (C-16), 109.7 (C-12), 103.9 (C-10), 90.6 (C-2), 82.0 (C-7), 62.5(C-1′), 61.6 (17-OCH₃), 61.1 (C-2′), 60.8 (C-3), 58.5 (C-21), 55.6(9-OCH₃), 51.3 (22-OCH₃), 50.1 (C-5), 40.4 (C-20), 40.2 (C-15), 35.3(C-6), 24.1 (C-14), 19.1 (C-19), 13.1 (C-18). CD (c=0.42 mM, MeOH, 24°C.), Δε (λ nm): 0 (310), −0.8 (287), −0.1 (262), −0.8 (247), 0 (241),+0.2 (236), 0 (229), −19.1 (212), −6.7 (200). EI-MS m/z (%): 494 (M⁺+2,25), 492 (M⁺, 65), 239 (100). HR-FAB-MS (NBA/PEG): calcd. for C₂₅H₃₄N₂O₆³⁵Cl: 493.2105, found: 493.2107, calcd. for C₂₅H₃₄N₂O₆ ³⁷Cl: 495.2089,found: 495.2082.

Bromination of Compound 3

To a solution of 3 (101 mg, 0.22 mmol) in dry DMF (2.0 mL) was added asolution of N-bromosuccinimide (43.2 mg, 0.24 mmol) in dry DMF (1.0 mL)and the mixture was stirred for 5 h at room temperature under argonatmosphere. The reaction mixture was poured into saturated aqueousNaHCO₃ solution, and this was extracted three times with chloroform. Thecombined extract was washed with brine, dried over MgSO₄, andconcentrated to give a residue that was purified by pre-packedamino-silica gel column chromatography (ethyl acetate/n-hexane=10:90) togive 90 mg (75%) of 13a and 29 mg (24%) of 13b, as a colorless amorphouspowder, respectively. Compound 13a; UV (MeOH) λ_(max) nm (log ε): 298(3.19), 281 (3.09), 245 (4.24), 228 (4.04), 211 (4.42). IR (ATR) ν_(max)cm⁻¹:2941, 1699, 1628, 1602, 1456, 1283. ¹H-NMR (500 MHz, CDCl₃) δ ppm:7.41 (1H, s, H-17), 7.27 (1H, d, J=8.2 Hz, H-11), 6.38 (1H, d, J=8.2 Hz,H-12), 4.33 (1H, br. s, N_(a)—H), 3.93 (3H, s, 9-OCH₃), 3.88 (2H, m,H-1′ and H-2′), 3.79 (3H, s, 17-OCH₃), 3.70 (3H, s, 22-OCH₃), 3.69 (1H,m, H-1′), 3.43 (1H, dd, J=11.6, 2.1 Hz, H-2′), 3.00 (1H, d, J=10.4 Hz,H-21), 2.92 (1H, ddd, J=13.4, 3.6, 3.6 Hz, H-15), 2.51 (1H, br. d,J=11.0 Hz, H-5), 2.45 (1H, br. d, J=10.4 Hz, H-3), 2.34 (2H, m, H-5 andH-14), 2.26 (1H, d, J=12.2 Hz, H-21), 2.16 (1H, ddd, J=14.3, 2.4, 2.4Hz, H-6), 1.89 (1H, ddd, J=13.7, 13.7, 4.0 Hz, H-6), 1.78 (1H, d, J=12.8Hz, H-14), 1.72 (1H, m, H-19), 1.57 (1H, br. d, J=11.3 Hz, H-20), 1.25(1H, m, H-19), 0.85 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz,CDCl₃) δ ppm: 169.0 (C-22), 160.4 (C-17), 153.8 (C-9), 148.7 (C-13),133.5 (C-11), 123.5 (C-8), 111.5 (C-16), 109.1 (C-12), 106.5 (C-10),91.1 (C-2), 81.5 (C-7), 62.7 (C-1′), 61.6 (17-OCH₃), 61.5 (9-OCH₃), 61.1(C-2′), 60.8 (C-3), 58.4 (C-21), 51.2 (22-OCH₃), 50.2 (C-5), 40.4(C-20), 39.9 (C-15), 36.5 (C-6), 24.0 (C-14), 19.1 (C-19), 13.0 (C-18).CD (c=0.19 mM, MeOH, 24° C.), Δε (λ nm): 0 (319), −0.9 (299), 0 (268),+0.3 (259), 0 (254), −17.2 (210), −8.7 (200). FAB-MS (NBA) m/z: 539[M+2+H]⁺, 537 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₅H₃₄N₂O₆ ⁷⁹Br:537.1600, found: 537.1615, calcd. for C₂₅H₃₄N₂O₆ ⁸¹Br: 539.1584, found:539.1589. Compound 13b; UV (MeOH) λ_(max) nm (log c): 294 (3.27), 275(3.02), 233 (4.16), 215 (4.54). IR (ATR) ν_(max) cm⁻¹: 2953, 1699, 1607,1484, 1278. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.42 (1H, s, H-17), 7.20 (1H,d, J=8.8 Hz, H-11), 6.32 (1H, d, J=8.8 Hz, H-10), 4.40 (1H, br. s,N_(a)—H), 3.92 (1H, ddd, J=11.6, 11.6, 3.1 Hz, H-2′), 3.83 (3H, s,9-OCH₃), 3.79 (3H, s, 17-OCH₃), 3.79 (1H, m, H-1′, overlapped with17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.68 (1H, dd, J=11.5, 2.7 Hz, H-1′),3.43 (1H, dd, J=11.6, 2.3 Hz, H-2′), 2.99 (1H, dd, J=11.6, 1.9 Hz,H-21), 2.92 (1H, ddd, J=12.9, 3.5, 3.5 Hz, H-15), 2.46 (2H, m, H-3 andH-5), 2.36 (2H, m, H-5 and H-14), 2.26 (1H, dd, J=11.7, 2.7 Hz, H-21),2.15 (1H, ddd, J=14.1, 2.4, 2.4 Hz, H-6), 1.82 (2H, m, H-6 and H-14),1.75 (1H, m, H-19), 1.57 (1H, br. d, J=10.8 Hz, H-20), 1.26 (1H, m,H-19), 0.85 (311, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δppm: 169.2 (C-22), 160.5 (C-17), 156.2 (C-9), 147.0 (C-13), 132.0(C-11), 117.3 (C-8), 111.6 (C-16), 104.5 (C-10), 97.4 (C-12), 90.4(C-2), 82.2 (C-7), 62.4 (C-1′), 61.6 (17-OCH₃), 61.1 (C-2′), 60.7 (C-3),58.4 (C-21), 55.6 (9-OCH₃), 51.3 (22-OCH₃), 50.1 (C-5), 40.4 (C-20),40.2 (C-15), 35.3 (C-6), 24.1 (C-14), 19.1 (C-19), 13.1 (C-18). CD(c=0.18 mM, MeOH, 24° C.), Δε (λ nm): 0 (308), −0.9 (290), +0.1 (258),−0.7 (250), 0 (248), +0.4 (229), 0 (225), −22.0 (212), −6.9 (200).FAB-MS (NBA) m/z: 539 [M+2+H]⁺, 537 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd.for C₂₅H₃₄N₂O₆ ⁷⁹Br: 537.1600, found: 537.1640, calcd. for C₂₅H₃₄N₂O₆⁸¹Br: 539.1584, found: 539.1642.

Nitration of Compound 3

To a solution of 3 (20.5 mg, 0.045 mmol) in dry DCM (0.9 mL) were addedcerium (IV) ammonium nitrate (24.4 mg, 0.045 mmol) and one drop of conc.H₂SO₄ and the mixture was stirred for 1 h at room temperature underargon atmosphere. The reaction mixture was poured into cold saturatedaqueous NaHCO₃ solution, and this was extracted three times withchloroform. The combined extract was washed with brine, dried overMgSO₄, and evaporated to give a residue that was purified by pre-packedamino-silica get column chromatography (ethylacetate/n-hexane=30:70-50:50) to give 11.6 mg (52%) of 14a and 4.7 mg(21%) of 14b, as a pale yellowish amorphous powder, respectively.Compound 14a; UV (MeOH) λ_(max) nm (log ε): 359 (3.87), 283 (3.29), 242(4.09), 228 (4.04), 206 (4.28). IR (ATR) ν_(max) cm⁻¹: 2949, 1698, 1599,1517, 1326, 1241. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.93 (1H, d, J=8.6 Hz,H-11), 7.42 (1H, s, H-17), 6.47 (1H, d, J=8.6 Hz, H-12), 4.87 (1H, br.s, N_(a)—H), 3.97 (3H, s, 9-OCH₃), 3.89 (2H, m, H-1′ and H-2′), 3.81(3H, s, 17-OCH₃), 3.73 (1H, m, H-1′, overlapped with 22-OCH₃), 3.71 (3H,s, 22-OCH₃), 3.46 (1H, d, J=9.7 Hz, H-2′), 3.00 (1H, dd, J=11.7, 1.8 Hz,H-21), 2.93 (1H, ddd, J=13.2, 3.7, 3.7 Hz, H-15), 2.52 (1H, d, J=10.4Hz, H-5), 2.47 (1H, d, J=9.3 Hz, H-3), 2.31 (3H, m, H-5, H-14 and H-21),2.21 (1H, d, J=14.3 Hz, H-6), 1.86 (1H, ddd, J=13.3, 13.3, 4.5 Hz, H-6),1.80 (1H, d, J=12.6 Hz, H-14), 1.70 (1H, m, H-19), 1.59 (1H, br. d,J=11.7 Hz, H-20), 1.26 (1H, m, H-19), 0.86 (3H, dd, J=7.3, 7.3 Hz,H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 168.9 (C-22), 160.4 (C-17),154.2 (C-13), 153.0 (C-9), 135.6 (C-10), 129.3 (C-11), 123.1 (C-8),111.5 (C-16), 106.3 (C-12), 91.0 (C-2), 81.0 (C-7), 62.8 (9-OCH₃), 62.6(C-1′), 61.7 (17-OCH₃), 61.0 (C-2′), 60.6 (C-3), 58.4 (C-21), 51.3(22-OCH₃), 50.0 (C-5), 40.3 (C-20), 39.8 (C-15), 36.2 (C-6), 24.0(C-14), 19.1 (C-19), 13.0 (C-18). CD (c=0.26 mM, MeOH, 24° C.), As (Anm): 0 (436), +1.4 (379), 0 (344), −1.0 (304), 0 (286), +0.2 (282), 0(278), −1.9 (263), 0 (249), +0.6 (242), 0 (237), −11.3 (208), −10.5(200). FAB-MS (NBA) m/z: 504 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₂₅H₃₄N₃O₈: 504.2346, found: 504.2318. Compound 14b; UV (MeOH) λ_(max)nm (log ε): 356 (3.66), 314 (3.84), 274 (3.38), 241 (4.23), 221 (4.02),205 (4.37). IR (ATR) ν_(max) cm⁻¹: 2944, 1699, 1625, 1596, 1476, 1324,1274, 1252. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 8.00 (1H, d, J=9.3 Hz, H-11),7.46 (1H, s, H-17), 6.87 (1H, br. s, N_(a)—H), 6.46 (1H, d, J=9.5 Hz,H-10), 3.95 (3H, s, 9-OCH₃), 3.91 (1H, m, H-2′, overlapped with 9-OCH₃),3.82 (3H, s, 17-OCH₃), 3.73 (2H, m, H₂-1′, overlapped with 22-OCH₃),3.72 (3H, s, 22-OCH₃), 3.46 (1H, d, J=11.0 Hz, H-2′), 2.99 (1H, dd,J=11.5, 1.8 Hz, H-21), 2.94 (1H, m, H-15), 2.47 (4H, m, H-3, H₂-5 andH-14), 2.28 (1H, dd, J=11.5, 2.9 Hz, H-21), 2.18 (1H, ddd, J=14.3, 2.3,2.3 Hz, H-6), 1.88 (1H, m, H-14), 1.79 (111, ddd, J=13.7, 13.7, 4.5 Hz,H-6), 1.72 (1H, m, H-19), 1.59 (1H, br. d, J=12.8 Hz, H-20), 1.26 (1H,m, H-19), 0.85 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃)δ ppm: 169.2 (C-22), 161.3 (C-9), 160.7 (C-17), 146.8 (C-13), 127.3(C-12), 126.9 (C-11), 117.3 (C-8), 111.1 (C-16), 103.5 (C-10), 90.0(C-2), 80.7 (C-7), 62.6 (C-1′), 61.6 (17-OCH₃), 61.0 (C-2′), 60.4 (C-3),58.2 (C-21), 56.1 (9-OCH₃), 51.3 (22-OCH₃), 49.8 (C-5), 40.2 (C-20),40.2 (C-15), 35.3 (C-6), 24.4 (C-14), 19.0 (C-19), 12.9 (C-18). CD(c=0.24 mM, MeOH, 24° C.), Δε (λ nm): 0 (437), +1.4 (392), 0 (363), −2.1(333), 0 (309), +1.8 (288), 0 (259), −8.0 (235), −4.2 (215), −7.4 (203),−7.1 (200). FAB-MS (NBA) m/z: 504 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd.for C₂₅H₃₄N₃O₈: 504.2346, found: 504.2338.

Introduction of a Methoxyl Group on C-10 in Compound 3

To a solution of 3 (50.8 mg, 0.11 mmol) in dry MeOH (2.0 mL) and dry DCM(0.5 mL) was added iodosobenzene diacetate (73.2 mg, 0.22 mmol) and themixture was stirred for 30 min at 0° C. under argon atmosphere. Thereaction mixture was poured into saturated aqueous NaHCO₃ solution, andthis was extracted three times with chloroform. The combined extract waswashed with brine, dried over MgSO₄, and evaporated to give a residuethat was purified by pre-packed silica gel column chromatography (2%MeOH in CHCl₃) to give 39.4 mg (69%) of an intermediate: UV (MeOH)λ_(max) nm: 296, 239, 220. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.43 (1H, s,H-17), 6.85 (1H, d, J=10.1 Hz, H-12), 6.16 (1H, d, J=10.1 Hz, H-11),4.08 (3H, s, 9-OCH₃), 3.92 (1H, m, H-2′), 3.77 (3H, s, 17-OCH₃), 3.73(2H, m, H₂-1′), 3.69 (3H, s, 22-OCH₃), 3.52 (1H, ddd, J=11.9, 2.3, 2.3Hz, H-2′), 3.26 (3H, s, 10-OCH₃), 3.24 (3H, s, 10-OCH₃), 2.95 (1H, dd,J=11.6, 1.8 Hz, H-21), 2.89 (1H, ddd, J=13.7, 3.1, 3.1 Hz, H-15), 2.84(1H, m, H-14), 2.54 (1H, dd, J=10.7, 2.7 Hz, H-3), 2.48 (1H, ddd,J=11.3, 4.0, 4.0 Hz, H-5), 2.28 (1H, ddd, J=11.7, 11.7, 2.8 Hz, H-5),2.20 (1H, dd, J=11.3, 3.1 Hz, H-21), 2.14 (1H, ddd, J=14.3, 3.1, 3.1 Hz,H-6), 1.89 (1H, br. d, J=13.4 Hz, H-14), 1.80 (1H, ddd, J=14.2, 11.5,4.6 Hz, H-6), 1.66 (1H, m, H-19), 1.53 (1H, br. d, J=11.3 Hz, H-20),1.24 (1H, m, H-19), 0.81 (3H, dd, J=7.5, 7.5 Hz, H₃-18). ¹³C-NMR (125MHz, CDCl₃) δ ppm: 169.7, 169.4 (C-13 and C-22), 160.6 (C-17), 150.9(C-9), 138.5 (C-11), 128.7 (C-12), 123.3 (C-8), 111.5 (C-16), 99.1(C-10), 95.3 (C-2), 79.4 (C-7), 63.2 (C-3), 62.5 (C-1′), 61.7 (17-OCH₃),60.8 (C-2′), 59.4 (9-OCH₃), 58.9 (C-21), 51.4 (10-OCH₃×₂), 51.2(22-OCH₃), 51.0 (C-5), 40.5 (C-20), 40.1 (C-15), 35.5 (C-6), 24.7(C-14), 19.1 (C-19), 13.0 (C-18). FAB-MS (NBA) m/z: 519 [M+H]⁺.

To a solution of the intermediate in dry MeOH (3.0 mL) was addedactivated Zn (445 mg) and the mixture was stirred for 85 h at refluxtemperature under argon atmosphere. The reaction mixture was filteredthrough Celite. The filtrate was concentrated under reduced pressure andpoured into saturated aq. NaHCO₃ solution, and this was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated. The crude product was chromatographed onamino-silica gel (ethyl acetate) to give 34.5 mg (64% in two steps) of15 as an amorphous powder. Compound 15; UV (MeOH) λ_(max) nm (log ε):305 (3.02), 281 (2.90), 241 (3.83), 226 (3.72), 207 (4.01). IR (KBr)ν_(max) cm⁻¹: 3327, 2948, 1703, 1627, 1487, 1261. ¹H-NMR (600 MHz,CDCl₃) δ ppm: 7.41 (1H, s, H-17), 6.70 (1H, d, J=8.2 Hz, H-11), 6.39(1H, d, J=8.2 Hz, H-12), 4.06 (1H, br. s, N_(a)—H), 3.93 (3H, s,9-OCH₃), 3.92 (2H, m, H-1′ and H-2′, overlapped with 9-OCH₃), 3.80 (3H,s, 10-OCH₃), 3.79 (3H, s, 17-OCH₃), 3.70 (3H, s, 22-OCH₃), 3.69 (1H, m,H-1′, overlapped with 22-OCH₃), 3.43 (1H, d, J=9.3 Hz, H-2′), 2.99 (1H,d, J=10.7 Hz, H-21), 2.91 (1H, d, J=13.2 Hz, H-15), 2.49 (1H, d, J=11.5Hz, H-5), 2.45 (1H, d, J=11.0 Hz, H-3), 2.34 (2H, m, H-5 and H-14), 2.25(1H, d, J=12.0 Hz, H-21), 2.14 (1H, d, J=14.4 Hz, H-6), 1.89 (1H, ddd,J=13.8, 13.8, 4.5 Hz, H-6), 1.77 (1H, d, J=11.3 Hz, H-14), 1.76 (1H, m,H-19), 1.57 (1H, br.d, J=11.3 Hz, H-20), 1.24 (1H, m, H-19), 0.84 (3H,dd, J=7.4, 7.4 Hz, H₃-18). ¹³C-NMR (150 MHz, CDCl₃) δ ppm: 169.2 (C-22),160.4 (C-17), 146.8 (C-9 and C-10), 142.1 (C-13), 122.7 (C-8), 113.7(C-11), 111.8 (C-16), 106.5 (C-12), 91.4 (C-2), 81.7 (C-7), 62.6 (C-1′),61.7 (17-OCH₃), 61.3 (C-2′), 61.0 (C-3 and 9-OCH₃), 58.7 (C-21), 56.7(10-OCH₃), 51.4 (22-OCH₃), 50.4 (C-5), 40.5 (C-20), 40.2 (C-15), 36.3(C-6), 24.1 (C-14), 19.2 (C-19), 13.1 (C-18). CD (c=0.21 mM, MeOH, 24°C.), Δε (λ nm): 0 (348), +0.5 (316), 0 (302), −0.7 (287), 0 (279), +3.4(256), 0 (245), −14.0 (208), −6.1 (200). FAB-MS (NBA) m/z: 489 [M+H]⁺.HR-FAB-MS (NBA/PEG): calcd. for C₂₆H₃₇N₂O₇: 489.2601, found: 489.2555.

Preparation of 10-Fluoromitragynine (17)

To a stirred solution of 11 (168 mg, 0.35 mmol) in dry AcOH (3.5 mL) wasadded NaCNBH₃ (46.7 mg, 0.71 mmol) at room temperature. After 15 min,another portion of NaCNBH₃ (46.7 mg, 0.71 mmol) was added to thereaction mixture, and this was stirred for 1 h at room temperature underargon atmosphere. MeOH (45 μL) was added to the reaction mixture, whichwas then heated under reflux at 70° C. for 7 h. The reaction mixture waspoured into cold aqueous NH₄OH solution, and this was extracted fourtimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated to give a residue that was purified bypre-packed amino-silica gel column chromatography (ethylacetate/n-hexane=30:70-50:50) to give 131.3 mg (89%) of 17 as acolorless amorphous powder. Compound 17; UV (MeOH) λ_(max) nm (log ε):275 (3.73), 227 (4.33), 209 (4.16). IR (KBr) ν_(max) cm⁻¹: 2952, 1700,1646, 1508, 1439, 1241. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.74 (1H, br. s,N_(a)—H), 7.44 (1H, s, H-17), 6.89 (1H, dd, J=8.5, 3.7 Hz, H-12), 6.84(1H, dd, J=11.6, 8.5 Hz, H-11), 3.99 (3H, d, J=1.2 Hz, 9-OCH₃), 3.73(3H, s, 17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.15 (1H, d, J=9.8 Hz, H-3),3.10 (1H, m, H-6), 3.03 (2H, m, H-15 and H-21), 2.93 (2H, m, H-5 andH-6), 2.56 (1H, dd, J=11.5, 4.5 Hz, H-5), 2.51 (1H, m, H-14), 2.45 (1H,dd, J=11.6, 2.4 Hz, H-21), 1.79 (1H, m, H-14), 1.76 (1H, m, H-19), 1.63(1H, br. d, J=11.0 Hz, H-20), 1.21 (1H, m, H-19), 0.87 (3H, dd, J=7.3,7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 169.2 (C-22), 160.6(C-17), 149.1 (d, J=233.7 Hz, C-10), 139.7 (d, J=12.4 Hz, C-9), 136.5(C-2), 133.6 (C-13), 121.7 (C-8), 111.2 (C-16), 110.4 (d, J=23.8 Hz,C-11), 107.6 (d, J=5.0 Hz, C-7), 105.9 (d, J=8.2 Hz, C-12), 62.0 (d,J=4.6 Hz, 9-OCH₃), 61.5 (17-OCH₃), 61.3 (C-3), 57.7 (C-21), 53.6 (C-5),51.3 (22-OCH₃), 40.5 (C-20), 39.7 (C-15), 29.7 (C-14), 23.3 (C-6), 19.0(C-19), 12.8 (C-18). CD (c=0.28 mM, MeOH, 24° C.), Δε (λ nm): 0 (324),+0.8 (300), +0.6 (294), +1.2 (274), 0 (259), −5.7 (240), −0.6 (229),−5.6 (215), +0.2 (200). FAB-MS (NBA) m/z: 417 [M+H]⁺. HR-FAB-MS(NBA/PEG): calcd. for C₂₃H₃₀N₂O₄F: 417.2190, found: 417.2192. FIG. 9 ais the 1H-NMR for Compound 17 and FIG. 9 b is the 13C-NMR for Compound17.

Preparation of 10-Chloromitragynine (18)

To a stirred solution of 12a (14.7 mg, 0.03 mmol) in dry AcOH (0.5 mL)was added NaCNBH₃ (4.0 mg, 0.06 mmol) at room temperature. After 15 min,another portion of NaCNBH₃ (4.0 mg, 0.06 mmol) was added to the reactionmixture, and this was stirred for 1 h at room temperature under argonatmosphere. MeOH (40 μL) was added to the reaction mixture, which wasthen heated under reflux at 90° C. for 18.5 h. The reaction mixture waspoured into cold aqueous NH₄OH solution, and this was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated to give a residue that was purified by silicagel column chromatography (ethyl acetate/n-hexane=50:50) to give 11.7 mg(91%) of 18 as a colorless amorphous powder. Compound 18; UV (MeOH)λ_(max) nm (log ε): 279 (3.64), 271 (3.63), 233 (4.40), 210 (4.16). IR(ATR) ν_(max) cm⁻¹: 2939, 1697, 1647, 1457, 1239. ¹H-NMR (500 MHz,CDCl₃) δ ppm: 7.88 (1H, br. s, N_(a)—H), 7.43 (1H, s, H-17), 7.04 (1H,d, J=8.2 Hz, H-11), 6.98 (1H, d, J=8.5 Hz, H-12), 3.93 (3H, s, 9-OCH₃),3.73 (3H, s, 17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.15 (1H, d, J=8.8 Hz,H-3), 3.10 (1H, m, H-6), 3.04 (1H, m, H-15), 3.03 (1H, m, H-21), 2.96(1H, m, H-5), 2.92 (1H, m, H-6), 2.55 (1H, m, H-5), 2.51 (1H, m, H-14),2.46 (1H, m, H-21), 1.80 (1H, d, J=13.1 Hz, H-14), 1.75 (1H, m, H-19),1.65 (1H, br. d, J=11.5 Hz, H-20), 1.23 (1H, m, H-19), 0.87 (3H, dd,J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 169.1 (C-22),160.5 (C-17), 149.1 (C-9), 136.4 (C-13*), 136.2 (C-2*), 122.6 (C-11),122.4 (C-8^(†)), 117.1 (C-10^(†)), 111.4 (C-16), 107.8 (C-12), 107.2(C-7), 61.9 (9-OCH₃), 61.6 (17-OCH₃), 61.2 (C-3), 57.8 (C-21), 53.6(C-5), 51.4 (22-OCH₃), 40.6 (C-20), 39.9 (C-15), 29.8 (C-14), 23.2(C-6), 19.1 (C-19), 12.8 (C-18), *^(,†): interchangeable. CD (c=0.15 mM,MeOH, 24° C.), Δε (λ nm): 0 (319), +1.0 (274), 0 (263), −7.1 (242), 0(234), +1.4 (229), 0 (224), −3.9 (215), 0 (204), +3.0 (200). FAB-MS(NBA) m/z: 435 [M+2+H]⁺, 433 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₂₃H₃₀N₂O₄ ³⁵Cl: 433.1894. found: 433.1856, calcd. for C₂₃H₃₀N₂O₄ ³⁷Cl:435.1875, found: 435.1853

Preparation of 10-Bromomitragynine (19)

To a stirred solution of 13a (12.1 mg, 0.023 mmol) in dry AcOH (0.4 mL)was added NaCNBH₃ (3.0 mg, 0.045 mmol) at room temperature. After 15min, another portion of NaCNBH₃ (3.0 mg, 0.045 mmol) was added to thereaction mixture, and this was stirred for 1 h at room temperature underargon atmosphere. MeOH (20 mL) was added to the reaction mixture, whichwas then heated under reflux at 90° C. for 12 h. The reaction mixturewas poured into cold aqueous NH₄OH solution, and this was extractedthree times with chloroform. The combined extract was washed with brine,dried over MgSO₄, and evaporated to give a residue that was purified bypre-packed amino-silica gel column chromatography (ethylacetate/n-hexane=30:70-50:50) to give 10.3 mg (96%) of 19 as a colorlessamorphous powder. Compound 19; UV (MeOH) λ_(max) nm (log ε): 279 (3.67),231 (4.47), 210 (4.21). IR (KBr) ν_(max) cm⁻¹: 2954, 1699, 1636, 1458,1246. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.90 (1H, br. s, N_(a)—H), 7.44(1H, s, H-17), 7.18 (1H, d, J=8.4 Hz, H-11), 6.95 (1H, d, J=8.4 Hz,H-12), 3.92 (3H, S, 9-OCH₃), 3.74 (3H, s, 17-OCH₃), 3.71 (3H, s,22-OCH₃), 3.15 (1H, d, J=11.5 Hz, H-3), 3.10 (1H, m, H-6), 3.03 (2H, m,H-15 and H-21), 2.95 (2H, m, H-5 and H-6), 2.56 (1H, m, H-5), 2.51 (1H,m, H-14), 2.46 (1H, dd, J=11.6, 2.7 Hz, H-21), 1.81 (1H, d, J=12.4 Hz,H-14), 1.75 (1H, m, H-19), 1.64 (1H, br. d, J=11.0 Hz, H-20), 1.22 (1H,m, H-19), 0.87 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (100 MHz, CDCl₃)δ ppm: 169.1 (C-22), 160.5 (C-17), 150.1 (C-9), 137.1 (C-13), 136.0(C-2), 125.2 (C-11), 122.6 (C-8), 111.3 (C-16), 108.4 (C-12), 107.1(C-7), 106.0 (C-10), 62.1 (9-OCH₃), 61.6 (17-OCH₃), 61.2 (C-3), 57.7(C-21), 53.6 (C-5), 51.4 (22-OCH₃), 40.6 (C-20), 39.9 (C-15), 29.7(C-14), 23.2 (C-6), 19.1 (C-19), 12.8 (C-18). CD (c=0.28 mM, MeOH, 24°C.), Δε (λ nm): 0 (324), +0.9 (275), 0 (263), −7.3 (243), 0 (234), +2.0(230), 0 (224), −3.8 (213), 0 (205), +3.6 (200). FAB-MS (NBA) m/z: 479[M+2+H]⁺, 477 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₃H₃₀N₂O₄ ⁷⁹Br:477.1389, found: 477.1393, calcd. for C₂₃H₃₀N₂O₄ ⁸¹Br: 479.1372, found:479.1377.

Preparation of 10-Nitromitragynine (19) via Indolenine 16

To a solution of 14a (56.8 mg, 0.11 mmol) in dry DCM (1.0 mL) were added2,6-lutidine (66 μL, 0.57 mmol) and tert-butyldimethylsilyltrifluoromethanesulfonate (66 μL, 0.28 mmol) and the mixture was stirredfor 23 h at room temperature under

argon atmosphere. The reaction mixture was poured into cold saturatedaqueous NaHCO₃ solution, and this was extracted three times withchloroform. The combined extract was washed with brine, dried overMgSO₄, and evaporated to give a residue that was purified by pre-packedsilica gel column chromatography (ethyl acetate/n-hexane=25:75) and thenamino-silica gel column chromatography (CHCl₃/n-hexane=30:70) to give60.6 mg (87%) of 16 as an amorphous powder. Compound 16; UV (MeOH)λ_(max) nm: 284, 275, 243 (sh), 214, 201. ¹H-NMR (400 MHz, CDCl₃) δ ppm:7.94 (1H, d, J=8.4 Hz, H-11), 7.45 (1H, s, H-17), 7.41 (1H, d, J=8.4 Hz,H-12), 4.04 (3H, s, 9-OCH₃), 3.83 (3H, s, 17-OCH₃), 3.71 (3H, s,22-OCH₃), 3.68 (2H, m), 3.05 (4H, m), 2.87 (3H, m), 2.76 (1H, d, J=13.9Hz), 2.63 (1H, d, J=11.5 Hz), 2.48 (1H, d, J=11.5 Hz), 1.84 (1H, d,J=13.7 Hz), 1.64 (3H, m), 1.25 (1H, m, H-19), 0.89 (9H, s,—OSi(CH₃)₂C(CH)₃), 0.83 (311, dd, J=7.3, 7.3 Hz, H₃-18), 0.07 (3H, s,—OSi(CH)₂C(CH₃)₃), 0.05 (3H, s, —OSi(CH₃)₂C(CH₃)₃. ¹³C-NMR (125 MHz,CDCl₃) δ ppm: 188.8 (C-2), 169.1 (C-22), 160.7 (C-17), 159.2 (C-13),150.9 (C-9), 141.9 (C-10), 130.8 (C-8), 128.3 (C-11), 116.8 (C-12),111.5 (C-16), 86.6 (C-7), 66.1 (C-1′), 62.6 (9-OCH₃), 62.2 (C-2′), 62.0(C-3), 61.8 (17-OCH₃), 58.1 (C-21), 51.3 (22-OCH₃), 50.0 (C-5), 40.4(C-20), 39.0 (C-15), 35.6 (C-6), 25.9 (—OSi(CH₃)₂C(CH₃)₃), 25.8 (C-14),18.9 (C-19), 18.4 (—OSi(CH₃)₂C(CH₃)₃), 12.8 (C-18), −5.2(—OSi(H₃)₂C(CH₃)₃), −5.4 (—OSi(CH₃)₂C(CH₃)₃). FAB-MS (NBA) m/z: 618[M+H]J.

To a stirred solution of 16 (95.9 mg, 0.16 mmol) in dry AcOH (2.0 mL)was added NaCNBH₃ (19.9 mg, 0.30 mmol) at room temperature. After 10min, another portion of NaCNBH₃ (19.9 mg, 0.30 mmol) was added to thereaction mixture, and this was stirred for 30 min at room temperatureunder argon atmosphere. MeOH (20 μL) was added to the reaction mixture,which was then heated under reflux at 90° C. for 13 h. The reactionmixture was poured into cold aqueous NH₄OH solution, and this wasextracted three times with chloroform. The combined extract was washedwith brine, dried over MgSO₄, and evaporated to give a residue that waspurified by silica gel column chromatography (ethylacetate/n-hexane=40:60-100:0) to give 65.0 mg (94%) of 20 as a colorlessamorphous powder. Compound 20; UV (MeOH) λ_(max) nm (log ε): 327 (3.71),306 (3.60), 260 (4.23), 239 (4.21), 217 (5.36). IR (KBr) ν_(max) cm⁻¹:3387, 2952, 1701, 1620, 1520, 1330, 1245. ¹H-NMR (400 MHz, CDCl₃) 5 ppm:8.07 (1H, br. s, N_(a)—H), 7.77 (1H, d, J=9.0 Hz, H-11), 7.44 (1H, s,H-17), 7.05 (1H, d, J=8.8 Hz, H-12), 4.02 (3H, s, 9-OCH₃), 3.75 (3H, s,17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.18 (1H, d, J=13.9 Hz, H-3), 3.16 (1H,m, H-6), 3.05 (2H, m, H-15 and H-21), 2.98 (2H, m, H-5 and H-6), 2.56(2H, m, H-5 and H-14), 2.48 (1H, d, J=9.2 Hz, H-21), 1.82 (1H, d, J=12.8Hz, H-14), 1.75 (1H, m, H-19), 1.65 (1H, br. d, J=13.2 Hz, H-20), 1.24(1H, m, H-19), 0.88 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz,CDCl₃) δ ppm: 169.0 (C-22), 160.6 (C-17), 149.9 (C-9), 139.9 (C-13),137.5 (C-2), 135.4 (C-10), 122.0 (C-8), 119.2 (C-11), 111.2 (C-16),109.9 (C-7), 106.8 (C-12), 107.1 (C-7), 63.5 (9-OCH₃), 61.6 (17-OCH₃),61.0 (C-3), 57.7 (C-21), 53.3 (C-5), 51.4 (22-OCH₃), 40.5 (C-20), 39.7(C-15), 29.7 (C-14), 23.3 (C-6), 19.1 (C-19), 12.8 (C-18). CD (c=0.28mM, MeOH, 24° C.), Δε (λ nm): 0 (410), −1.4 (357), 0 (325), +5.3 (273),0 (261), −6.9 (237), +0.1 (226), +2.6 (221), −0.1 (217), −7.8 (206),−5.3 (200). FAB-MS (NBA) m/z: 444 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd.for C₂₃H₃₀N₃O₆: 444.2135, found: 444.2104.

Preparation of 10-Methoxymitragynine (21)

To a stirred solution of 15 (41.2 mg, 0.084 mmol) in dry AcOH (1.0 mL)was added NaCNBH₃ (11.2 mg, 0.17 mmol) at room temperature. After 15min, another portion of NaCNBH₃ (11.2 mg, 0.17 mmol) was added to thereaction mixture, and this was stirred for 1 h at room temperature underargon atmosphere. MeOH (40 mL) was added to the reaction mixture, whichwas then heated under reflux at 90° C. for 5 h. The reaction mixture waspoured into cold aqueous NH₄OH solution, and this was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated to give a residue that was purified bypre-packed amino-silica gel column chromatography (ethylacetate/n-hexane=30:70) and then by silica gel column chromatography(ethyl acetate/n-hexane=40:60) to give 23.2 mg (64%) of 21 as acolorless amorphous powder. Compound 21; UV (MeOH) λ_(max) nm (log ε):276 (3.76), 229 (4.40), 208 (4.27). IR (KBr) ν_(max) cm⁻¹: 3358, 2936,1702, 1643, 1502, 1435, 1244. ¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.58 (1H,br s, N_(a)—H), 7.43 (1H, s, H-17), 6.95 (1H, d, J=8.6 Hz, H-11), 6.81(1H, d, J=8.8 Hz, H-12), 3.92 (3H, s, 9-OCH₃), 3.88 (3H, s, 10-OCH₃),3.73 (3H, s, 17-OCH₃), 3.71 (3H, s, 22-OCH₃), 3.15 (1H, m, H-3), 3.13(1H, m, H-6), 3.02 (2H, m, H-15 and H-21), 2.95 (1H, m, H-5 and H-6),2.55 (1H, m, H-5), 2.50 (1H, m, H-14), 2.45 (1H, m, H-21), 1.79 (11, d,−14.6 Hz, H-14), 1.75 (1H, m, H-19), 1.63 (1H, br. d, J=10.6 Hz, H-20),1.21 (1H, m, H-19), 0.87 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (100MHz, CDCl₃) δ ppm: 169.2 (C-22), 160.5 (C-17), 145.5 (C-10), 142.6(C-9), 136.1 (C-2), 133.2 (C-13), 122.3 (C-8), 111.5 (C-16), 110.6(C-11), 107.1 (C-7), 106.0 (C-12), 61.7 (17-OCH₃), 61.5 (9-OCH₃), 61.3(C-3), 58.2 (10-OCH₃), 57.8 (C-21), 53.7 (C-5), 51.3 (22-OCH₃), 40.6(C-20), 39.9 (C-15), 29.8 (C-14), 23.4 (C-6), 19.1 (C-19), 12.8 (C-18).CD (c=0.31 mM, MeOH, 24° C.), Δε (λ nm): 0 (323), +0.1 (318), 0 (315),−0.1 (307), 0 (304), +2.2 (274), 0 (260), −6.3 (243), −2.6 (229), −5.9(218), −5.8 (216), −6.3 (212), 0 (204), +0.8 (200). FAB-MS (NBA) m/z:429 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₄H₃₃N₂O₅: 429.2389, found:429.2376.

Preparation of 7-Hydroxy-10-fluoromitragynine (22)

A solution of PIFA (56.0 mg, 0.13 mmol) in MeCN (0.5 mL) was addeddropwise to a stirred solution of 17 (51.8 mg, 0.12 mmol) in MeCN (1.7mL) and H₂O (0.6 mL) at 0° C. and the mixture was stirred for 2 h at thesame temperature under argon atmosphere. The reaction mixture was pouredinto cold saturated aqueous NaHCO₃ solution, and this was extractedthree times with chloroform. The combined extract was washed with brine,dried over MgSO₄, and concentrated to give a residue that was purifiedby pre-packed amino-silica gel column chromatography (ethylacetate/n-hexane=30:70-50:50) to give 30.2 mg (56%) of 22 as a colorlessamorphous powder. Compound 22; UV (MeOH) λ_(max) nm (log ε): 295 (3.37),287 (3.37), 245 (sh, 4.03), 221 (4.22), 207 (4.13), 201 (4.15). IR (KBr)ν_(max) cm⁻¹: 3395, 2952, 1702, 1645, 1488, 1250. ¹H-NMR (500 MHz,CDCl₃) δ ppm: 7.43 (1H, s, H-17), 7.18 (1H, dd, J=8.4, 3.6 Hz, H-12),7.04 (1H, dd, J=12.5, 8.2 Hz, H-11), 4.04 (3H, d, J=2.7 Hz, 9-OCH₃),3.80 (3H, s, 17-OCH₃), 3.67 (3H, s, 22-OCH₃), 3.12 (1H, dd, J=11.0, 2.4Hz, H-3), 3.04 (1H, dd, J=11.4, 3.1 Hz, H-21), 3.00 (1H, ddd, J=14.0,3.6, 3.6 Hz, H-15), 2.80 (2H, m, H-5 and H-14), 2.65 (1H, m, H-5), 2.60(1H, m, H-6), 2.49 (1H, dd, J=611.6, 2.7 Hz, H-21), 1.85 (1H, d, J=13.7Hz, H-14), 1.70 (2H, m, H-6 and H-19), 1.59 (1H, br.d, J=11.0 Hz, H-20),1.24 (1H, m, H-19), 0.82 (3H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125MHz, CDCl₃) δ ppm: 183.9 (C-2), 169.3 (C-22), 160.8 (C-17), 153.6 (d,J=247.0 Hz, C-10), 150.2 (C-13), 144.0 (d, J=12.8 Hz, C-9), 132.3 (d,J=3.7 Hz, C-8), 117.5 (d, J=21.5 Hz, C-11), 115.8 (d, J=7.3 Hz, C-12),111.1 (C-16), 81.4 (C-7), 61.8 (17-OCH₃), 61.5 (d, J=4.6 Hz, 9-OCH₃),61.5 (C-3), 58.1 (C-21), 51.3 (22-OCH₃), 49.9 (C-5), 40.4 (C-20), 39.2(C-15), 35.9 (C-6), 26.0 (C-14), 18.9 (C-19), 12.8 (C-18). CD (c=0.27mM, MeOH, 24° C.), Δε (λ nm): 0 (341), +3.6 (299), +1.4 (280), +9.0(254), −0.1 (239), −11.8 (225), 0 (210), +9.0 (200). FAB-MS (NBA) m/z:433 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₃H₃₀N₂O₅F: 433.2139,found: 433.2140. FIG. 10 a is the ¹H-NMR for Compound 22 and FIG. 10 bis the ¹³C-NMR for Compound 22.

Preparation of 7-Hydroxy-10-chloromitragynine (23)

A solution of PIFA (26.1 mg, 0.059 mmol) in MeCN (0.2 mL) was addeddropwise to a stirred solution of 18 (23.1 mg, 0.053 mmol) in MeCN (0.9mL) and H₂O (0.3 mL) at 0° C. and the mixture was stirred at the sametemperature under argon atmosphere. After 1 h, another portion of PIFA(2.3 mg, 0.005 mmol) was added to the reaction mixture, and this wasfurther stirred for 3.5 h at 0° C. The reaction mixture was poured intocold saturated aqueous NaHCO₃ solution, and this was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated to give a residue that was purified by silicagel column chromatography (ethyl acetate/n-hexane=40:60) to give 12.8 mg(54%) of 23 as a colorless amorphous powder. Compound 23; UV (MeOH)λ_(max) nm (log ε): 303 (3.49), 244 (sh, 4.09), 224 (4.25), 209 (4.20),202 (4.22). IR (KBr) μ_(max) cm⁻¹: 3410, 2930, 1682, 1642, 1592, 1464,1247. ¹H-NMR (500 MHz, CDCl₃) δ ppm: 7.44 (14, s, H-17), 7.35 (1H, d,J=7.9 Hz, H-11), 7.26 (1H, d, J=7.9 Hz, H-12), 3.99 (3H, s, 9-OCH₃),3.81 (3H, s, 17-OCH₃), 3.66 (3H, s, 22-OCH₃), 3.13 (1H, dd, J=11.0, 2.4Hz, H-3), 3.05 (111, dd, J=1.3, 2.1 Hz, H-21), 3.00 (1H, ddd, J=14.0,3.7, 3.7 Hz, H-15), 2.79 (2H, m, H-5 and H-14), 2.67 (1H, m, H-5), 2.63(1H, m, H-6), 2.49 (1H, dd, J=11.1, 2.3 Hz, H-21), 1.85 (1H, d, J=13.7Hz, H-14), 1.69 (2H, m, H-6 and H-19), 1.59 (1H, br. d, J=11.0 Hz,H-20), 1.24 (1H, m, H-19), 0.82 (34, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR(125 MHz, CDCl₃) δ ppm: 184.4 (C-2), 169.3 (C-22), 160.8 (C-17), 153.9(C-9), 152.5 (C-13), 133.7 (C-8), 131.5 (C-11), 125.5 (C-10), 117.8(C-12), 111.1 (C-16), 81.4 (C-7), 62.1 (9-OCH₃), 61.8 (17-OCH₃), 61.5(C-3), 58.1 (C-21), 51.3 (22-OCH₃), 50.0 (C-5), 40.4 (C-20), 39.2(C-15), 36.0 (C-6), 26.0 (C-14), 18.9 (C-19), 12.8 (C-18). CD (c=0.32mM, MeOH, 24° C.), Δε (λ nm): 0 (346), +3.8 (303), +1.4 (282), +9.9(257), −0.1 (241), −13.2 (227), −0.1 (212), +8.5 (204), +5.9 (200).FAB-MS (NBA) m/z: 451 [M+2+H]⁺, 449 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd.for C₂₃H₃₀N₂O₅ ³⁵Cl: 449.1843, found: 449.1853, calcd. for C₂₃H₃₀N₂O₅³⁷Cl: 451.1825, found: 451.1775.

Preparation of 7-Hydroxy-10-bromomitragynine (24)

A solution of PIFA (12.9 mg, 0.029 mmol) in MeCN (0.2 mL) was addeddropwise to a stirred solution of 19 (13.9 mg, 0.029 mmol) in MeCN (0.5mL) and H₂O (0.2 mL) at 0° C. and the mixture was stirred at the sametemperature under argon atmosphere. After 1.5 h, another portion of PIFA(1.3 mg, 0.003 mmol) was added to the reaction mixture, and this wasfurther stirred for 3.5 h at 0° C. The reaction mixture was poured intocold saturated aqueous NaHCO₃ solution, and this was extracted threetimes with chloroform. The combined extract was washed with brine, driedover MgSO₄, and evaporated to

give a residue that was purified by pre-packed silica gel columnchromatography (ethyl acetate/n-hexane=40:60) to give 4.8 mg (33%) of 24as a colorless amorphous powder, together with 0.6 mg (6%) of recoveredstarting material 19. Compound 24; UV (MeOH) λ_(max) nm (log ε): 306(3.54), 248 (sh, 4.09), 221 (4.26), 208 (4.21), 201 (4.25). IR (KBr)ν_(max) cm⁻¹: 3393, 2935, 1670, 1646, 1459, 1249. ¹H-NMR (400 MHz,CDCl₃) δ ppm: 7.44 (1H, s, H-17), 7.53 (11, d, J=8.2 Hz, H-11), 7.21(1H, d, J=8.2 Hz, H-12), 3.99 (3H, s, 9-OCH₃), 3.81 (3H, s, 17-OCH₃),3.65 (3H, s, 22-OCH₃), 3.12 (1H, dd, J=11.2, 2.4 Hz, H-3), 3.05 (1H, dd,J=11.5, 2.0 Hz, H-21), 2.99 (1H, ddd, J=13.9, 3.5, 3.5 Hz, H-15), 2.79(2H, m, H-5 and H-14), 2.66 (1H, m, H-5), 2.63 (1H, d, J=15.2 Hz, H-6),2.49 (1H, dd, J=11.4, 3.0 Hz, H-21), 1.87 (1H, d, J=13.7 Hz, H-14), 1.69(2H, m, H-6 and H-19), 1.59 (1H, br. d, J=11.5 Hz, H-20), 1.24 (1H, m,H-19), 0.82 (13H, dd, J=7.3, 7.3 Hz, H₃-18). ¹³C-NMR (125 MHz, CDCl₃) δppm: 184.3 (C-2), 169.3 (C-22), 160.8 (C-17), 154.8 (C-9), 153.6 (C-13),133.8 (C-8), 134.6 (C-11), 118.4 (C-12), 114.8 (C-10), 111.1 (C-16),81.5 (C-7), 62.3 (9-OCH₃), 61.8 (17-OCH₃), 61.5 (C-3), 58.1 (C-21), 51.3(22-OCH₃), 50.0 (C-5), 40.4 (C-20), 39.2 (C-15), 36.0 (C-6), 26.0(C-14), 18.9 (C-19), 12.8 (C-18). CD (c=0.21 mM, MeOH, 24° C.), Δε (λnm): 0 (346), +4.2 (304), +1.4 (282), +10.9 (258), 0 (242), −14.0 (228),0 (214), +9.1 (205), +8.5 (200). FAB-MS (NBA) m/z: 495 [M+2+H]⁺, 493[M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. for C₂₃H₃₀N₂O₅ ⁷⁹Br: 493.1338,found: 493.1317, calcd. for C₂₃H₃₀N₂O₅ ⁸¹Br: 495.1321, found: 495.1375.

Preparation of 7-Hydroxy-10-methoxymitragynine (25)

A solution of PIFA (22.7 mg, 0.051 mmol) in MeCN (1.0 mL) was addeddropwise to a stirred solution of 21 (21.5 mg, 0.050 mmol) in MeCN (3.0mL) and H₂O (1.0 mL) at 0° C. and the mixture was stirred for 1 h at thesame temperature under argon atmosphere. The reaction mixture was pouredinto cold saturated aqueous NaHCO₃ solution, and this was extractedthree times with chloroform. The combined extract was washed with brine,dried over MgSO₄, and concentrated to give a residue that was purifiedby amino-silica gel column chromatography (ethyl acetate/n-hexane=30:70)to give 4.4 mg (20%) of 25 as a colorless amorphous powder. Compound 25;UV (MeOH)

λ_(max) nm (log ε): 311 (3.48), 297 (3.43), 236 (4.08), 215 (4.04), 202(4.12). IR (KBr) ν_(max) cm⁻¹: 3410, 2952, 1702, 1644, 1488, 1433, 1259.¹H-NMR (400 MHz, CDCl₃) δ ppm: 7.44 (1H, s, H-17), 7.24 (1H, d, J=8.2Hz, H-12*), 6.84 (1H, d, J=8.4 Hz, H-11*), 3.95 (3H, s, 9-OCH₃), 3.86(3H, s, 10-OCH₃), 3.80 (3H, s, 17-OCH₃), 3.69 (3H, s, 22-OCH₃), 3.11(1H, dd, J=11.2, 2.6 Hz, H-3), 3.02 (2H, m, H-15 and H-21), 2.79 (2H, m,H-5 and H-14), 2.62 (2H, m, H-5 and H-6), 2.47 (1H, dd, J=11.5, 2.7 Hz,H-21), 2.22 (1H, br. s, 7-OH), 1.86 (1H, d, J=13.7 Hz, H-14), 1.75 (1H,ddd, J=13.5, 13.5, 4.6 Hz, H-6), 1.71 (1H, m, H-19), 1.59 (1H, br. d,J=11.4 Hz, H-20), 1.25 (1H, m, H-19), 0.82 (3H, dd, J=7.3, 7.3 Hz,H₃-18), *: interchangeable. ¹³C-NMR (125 MHz, CDCl₃) δ ppm: 182.0 (C-2),169.3 (C-22), 160.7 (C-17), 151.5 (C-13), 147.9 (C-9), 145.9 (C-10),132.6 (C-8), 116.3 (C-11), 112.7 (C-12), 111.3 (C-16), 81.2 (C-7), 61.8(17-OCH₃), 61.5 (C-3 and 9-OCH₃), 58.2 (C-21), 56.2 (10-OCH₃), 51.3(22-OCH₃), 50.1 (C-5), 40.5 (C-20), 39.3 (C-15), 36.3 (C-6), 26.1(C-14), 19.0 (C-19), 12.8 (C-18). CD (c=0.25 mM, MeOH, 24° C.), Δε (λnm): 0 (334), +2.3 (316), +2.4 (306), 0 (289), −0.3 (285), 0 (281), +6.4(264), +7.6 (256), 0 (244), −9.6 (233), −9.3 (228), 0 (211), +4.0 (200).FAB-MS (NBA) m/z: 445 [M+H]⁺. HR-FAB-MS (NBA/PEG): calcd. forC₂₄H₃₃N₂O₆: 445.2339, found: 445.2314.

Magnus Assay Using Guinea Pig Ileum Preparations

Male albino guinea pigs (Japan SLC) weighing 320-550 g were used.Animals were housed in a temperature-controlled room at 24° C. withlights on from 07:00-19:00 and had free access to food and water. Theguinea-pig ileum was dissected and placed in Krebs-Henseleit solution(in mM: NaCl, 112.08; KCl, 5.90; CaCl₂, 1.97; MgCl₂, 1.18; NaH₂PO₄,1.22; NaHCO₃, 25.00, and glucose, 11.49). The ileum was set up under 1 gof tension in 5 mL of an organ bath containing the nutrient solution.The bath was maintained at 37° C. and continuously bubbled with a gasmixture of 95% O₂ and 5% CO₂. At the start of each experiment, themaximum response to acetylcholine (3 μM) was obtained for each tissue tocheck its suitability. Tissues were stimulated through platinumneedle-ring (a ring was placed 20 mm above the base of a needle 5 mm inlength) electrodes using square wave pulses of supramaximal voltage. Theileum was transmurally stimulated with monophasic pulses (0.2 Hz) at 0.3ms duration by a stimulator (SEN-7203, Nihon Kohden, Tokyo, Japan).Contractions were isotonically recorded with a displacement transducer(NEC, San-ei Instruments Ltd., Type 45347), a DC strain amplifier(San-ei 6M92), and a DC recorder (Hitachi, Mod 056, Tokyo, Japan). Allconcentration-response curves were constructed in a cumulative manner.The height of the twitch response to transmural stimulation was measuredbefore and after drug challenge. The twitch response remaining aftereach sample addition was determined by dividing the height after itsaddition by the height before addition multiplied by 100. To obtain thepercentage inhibition, this value was subtracted from 100. Agonistactivity was expressed as pD₂ value, which is the negative logarithm ofthe molar concentration required to produce 50% of the maximum responseto the drug (EC₅₀). To investigate the involvement of opioid receptorsin the inhibitory effect of the samples, the antagonistic effect of theopioid receptor antagonist naloxone was examined. The inhibitory effectsof samples on electrically stimulated contraction were regarded asopioid activities when the antagonistic effect of naloxone was observed.

1. An indole alkaloid compound having an opioid receptor agonisticeffect having the formula (a):

wherein X represents halogen atom or hydrogen, Q and A eachindependently represents hydrogen, alkyl, or aryl, E represents hydrogenor a hydroxyl group, m is 0 or 1, n is 0 or 1, q is 0 or 1, R₁ and R₂independently are hydrogen, hydroxy, alkoxy, aryloxy, alkyl, aryl,aralkyl, alkaryl, alkyl amide, amino, alkylamino, halogen, fluorinatedalkyl, fluorinated alkoxy, nitro group, or cyano group, R₃ represents anester group or carboxyl group, wherein the dashed lines each indicatesan optional double bond with the provisos that when q is 0 and X ishalogen then a 2,7 double bond is present and when m is 0, q is 1 and Xis halogen then a 1,2 double bond is present.
 2. The indole alkaloidcompound of claim 1, wherein R₁ and R₂ independently are C₁-C₆ alkoxy,R₃ represents an C₁-C₆ alkyl carboxylic acid ester group, and Q and A,if present, are hydrogen.
 3. A C10-halogenated indole alkaloid compoundhaving opioid receptor agonistic effect.
 4. The C10-halogenated indolealkaloid compound having the formulae:

wherein R is a halogen atom, R₁ and R₂ independently are hydrogen,hydroxy, alkoxy, aryloxy, alkyl, aryl, aralkyl, alkaryl, alkyl amide,amino, alkylamino, halogen, fluorinated alkyl, fluorinated alkoxy, nitrogroup, or cyano group, and R₃ represents an ester group or carboxylgroup.
 5. The C10-halogenated indole alkaloid compound of claim 4selected from the formulae:

wherein R is fluorine, chlorine, bromine, or iodine.
 6. TheC10-halogenated indole alkaloid compound of claim 4, whereinC10-halogenated indole alkaloid compound is C10-halogenated mitragynine,7-hydroxy-C10-halogenatedmitragynine, C10-halogenated2,3-dimethylindole, C10-halogenated tetrahydrocarbazole, C10-halogenatedindoloquinolizidine corynantheol, C10-halogenated dihydrocorynantheol,or C10-halogenated yohimbine, singly or in any combination thereof. 7.The C10-halogenated indole alkaloid compound of claim 4, wherein saidcompound is 10-halomitragynine.
 8. The C10-halogenated indole alkaloidcompound of claim 4, wherein said compound is7-hydroxy-10-halomitragynine.
 9. An indole alkaloid compound having anopioid receptor agonistic effect having the formula:

where X is hydrogen or a halogen atom, R₁ and R₂ independently arehydrogen, hydroxy, alkoxy, aryloxy, alkyl, aryl, aralkyl, alkaryl, alkylamide, amino, alkylamino, halogen, fluorinated alkyl, fluorinatedalkoxy, nitro group, or cyano group, and R₃ represents an ester group orcarboxyl group.
 10. The indole alkaloid compound of claim 9 having theformula:


11. A reaction product containing said compound of claim 1 in aconcentration of from about 5 wt % to 100 wt % of said compound.
 12. Apharmaceutical composition comprising a therapeutic effective amount ofan indole alkaloid compound of claim 1 or a pharmaceutically acceptablesalt thereof.
 13. The pharmaceutical composition of claim 12 furthercomprising a pharmaceutically acceptable carrier.
 14. The pharmaceuticalcomposition of claim 12 in a pharmaceutically deliverable form selectedfrom the group consisting of powder, tablet pill, capsule, solution,suspension, emulsion, cream, gel, and aerosol.
 15. The pharmaceuticalcomposition of claim 12, containing 0.1% to 100% of said indole alkaloidcompound or a pharmaceutically acceptable salt thereof.
 16. Apharmaceutical composition comprising a therapeutic amount of a compoundof claim 4 or a pharmaceutically acceptable salt thereof.
 17. Apharmaceutical composition comprising a therapeutic amount of a compoundof claim 9 or a pharmaceutically acceptable salt thereof.
 18. Thepharmaceutical composition of claim 12 in a pharmaceutically deliverableform selected from the group consisting of solutions, suspensions,emulsions, tablets, pills, pellets, granules, lozenges, capsules,capsules containing liquids, powders, sustained-release formulations,syrups, elixirs, creams, gels, suppositories, emulsions, aerosols, andsprays.
 19. A pharmaceutical composition containing 0.1% to 100% of saidindole alkaloid compound of claim 1 or a pharmaceutically acceptablesalt thereof.
 20. A method for eliciting a therapeutic effect in apatient in need thereof, comprising the step of administering to saidpatient an effective dose of an indole alkaloid compound of claim 1 or apharmaceutically acceptable salt thereof.
 21. The method of claim 21wherein said compound is selected from the formulae:

wherein R is fluorine, chlorine, bromine, or iodine, or

where X is hydrogen, fluorine, chlorine, bromine, or iodine.
 22. Themethod of claim 20 wherein said therapeutic effect comprises ananalgesic effect.
 23. The method of claim 20 wherein said step ofadministering is performed by a delivery route selected from the groupconsisting of oral, transdermal, inhalation, injection, infusion, andsuppository.
 24. The method of claim 20 wherein said patient is ananimal.
 25. The method of claim 20 wherein said patient is a mammal. 26.The method of claim 20 wherein said patient is a human.
 27. The methodof claim 20 wherein patient is a human and the therapeutic effect ispain treatment, where an effective total daily dosage of said compoundranges from about 0.1 mg to about 1,000 mg active compound/kg bodyweight of said patient.
 28. A method for making a C10-halogenated indolealkaloid derivative compound, comprising the steps of: reacting aCorynanthe-type indole alkaloid with hypervalent iodine in the presenceof ethylene glycol effective to provide a 2,3-ethylene glycol bridgedindoline adduct, effective to mask a 2, 3-7c bond of an indole nucleusof said adduct; introducing a halogen atom at a C10 position of theadduct via electrophilic aromatic substitution, providing aC10-halogenated adduct derivative; converting said C10-halogenatedadduct derivative into a corresponding C10-halogenated mitragyninederivative by reduction reaction effective to eliminate the ethyleneglycol bridge; optionally, converting said C10-halogenated mitragyninederivative into a corresponding 7-hydroxy-10-halomitragynine derivativeby oxidation reaction, and optionally isolating or otherwiseconcentrating said derivative.
 29. A method for making an indolealkaloid compound comprising reacting 7-hydroxymitragynine and/or7-hydroxy-10-halomitragynine with a reducing agent, such as NaBH₄ inMeOH, to yield a compound according to claim 9, and optionally isolatingor otherwise concentrating said compound.